IVAX : Web-Based Vaccine Design Platform

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iVAX is a web-based vaccine design platform that allows you to build an epitope driven vaccine directly from a pathogen genome in a matter or hours or days. Analyze pathogen gene sequences for conserved antigens to provide better strain coverage, build sequences of the most effective, immunogenic epitopes, optimize protein sequences for efficient, targeted immune protection. In addition to designing vaccines from the ground up you can also use iVAX to predict the efficacy of existing vaccines or vaccines in development. iVAX has been validated in producing 5 prototype vaccines : tularemia, vaccinia / smallpox, H. pylori, pandemic influenza H7N9 (bird flu) and pandemic influenza H1N1 (swine flu).

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  • Combination of ISPRI platform, homology analysis, protein engineering analytics and lab-based validation techniquesOne or multiple unique pathogen genome(s) -> epitope-driven vaccine Demonstrated in multiple tests on different pathogens
  • Unfortunately, in many cases, the haystack is bigger than we thought. The more isolates from a single species we sequence, the more we find how different one is from the next. For those pathogens where strain variation occurs, focusing on core genome may be a good solution for choosing vaccine antigens. Caveat – Strain specific genes may be associated with virulence and might want to consider those.
  • Not only do we need to think about the pathogen in selecting antigens. We also need to think about the host. We have always considered it to be important, if possible, to exclude vaccine components that are conserved with self. The rationale for avoiding cross-reactivity is self-evident.In addition, we’re now excluding sequences that are conserved with Human microbiota… can be seen as an extension of the self
  • Cross reactivity of putative epitopes is not constrained to exact sequence matching as the TCR only interacts with five of 9 peptides in the epitope sequence. To improve our prediction of homology we developed JanusMatrix, which takes into account the “two-faced” nature of epitopes. We can discover sequences that interact with pre-existing non-naive T cells by searching for homologs at MHC binding residues where TCR-facing residues are strictly conserved. This may be useful for avoiding T cell epitopes that are cross reactive with pre-existing T cells.
  • 2) T effector epitopes:HCV_G1_2440HCV_G1_2879HCV_G1_2485HCV_G1_NS4B_1769HCV_G1_NS4B_1725HCV_G1_ENV_255HCV_G1_NS4B_1790HCV_G1_NS2_732HCV_G1_2840HCV_G1_1605HCV_G1_2941HCV_G1_ENV_359HCV_G1_NS2_909HCV_G1_NS4B_1876HCV_G1_NS4b_1798HCV_G1_DEXDC_1246HCV_G1_NS4B_1910HCV_G1_1941HCV_G1_2898HCV_G1_NS5A_1988HCV_G1_2913HCV_G1_NS2_748You need to zoom in to see the name of the cluster.The legend is:- Light blue diamond (source): cluster from HCV.- Gray square: Predicted 9-mer epitope from the HCV cluster. - Blue triangle: 9-mer from human genome (Plasma proteome database), 100% TCR face identical to cluster-derived epitope.- Lavender circle: Human protein where cross-reactive (with HCV cluster) epitopes are present.
  • Vaccine. 2009 Mar 18;27(13):1889-97. doi: 10.1016/j.vaccine.2009.01.116. Epub 2009 Feb 2.A phase I clinical trial of a PER.C6 cell grown influenza H7 virus vaccine.Cox RJ, Madhun AS, Hauge S, Sjursen H, Major D, Kuhne M, Höschler K, Saville M, Vogel FR, Barclay W, Donatelli I, Zambon M, Wood J, Haaheim LR.SourceInfluenza Centre, The Gade Institute, University of Bergen, N-5021 Bergen, Norway. rebecca.cox@gades.uib.noAbstractAvian influenza H7 viruses have transmitted from poultry to man causing human illness and fatality, highlighting the need for pandemic preparedness against this subtype. We have developed and tested the first cell-based human vaccine against H7 avian influenza virus in a phase I clinical trial. Sixty healthy volunteers were intramuscularly vaccinated with two doses of split H7N1 virus vaccine containing 12 microg or 24 microghaemagglutinin alone or with aluminium hydroxide adjuvant (300 microg or 600 microg, respectively). The vaccine was well tolerated in all subjects and no serious adverse events occurred. The vaccine elicited low haemagglutination inhibition and microneutralisationtitres, although the addition of aluminium adjuvant augmented the antibody response. We found a higher number of antibody secreting cells and an association with IL-2 production in subjects with antibody response. In conclusion, our study shows that producing effective H7 pandemic vaccines is as challenging as has been observed for H5 vaccines.
  • http://www.novartis.com/newsroom/media-releases/en/2013/1743124.shtml
  • IVAX : Web-Based Vaccine Design Platform

    1. 1. iVAX Overview 1
    2. 2. iVAX Toolkit – Online Access
    3. 3. Design next-gen vaccines better/safer/faster Better understanding of vaccine MOA Whole (live/killed) vaccines Subunit vaccines GenomeDerived, Epitope Driven (GD-ED) Vaccines Improve vaccine safety and efficacy Accelerate Vaccine Design
    4. 4. T cells = Immune System Body Armor T cell response cannot prevent Infection but . . . T cell response can arm against Disease
    5. 5. Vaccine Design Tools and Techniques Analysis Engineering Validation 5
    6. 6. Comparative Genomics Impacts Vaccine Immunogen Selection Strain 1 dispensable genes core genome Strain 3 Strain 2 pangenome strain-specific genes
    7. 7. Epitope Cross-Reactivity Impacts Vaccine Immunogen Selection
    8. 8. EpiMatrix • EpiVax uses EpiMatrix to predict epitopes – matrix based prediction algorithm • Can predict either class I or class II MHC binding – MHC binding is a prerequisite for immunogenicity Protein MHC II Pocket Peptide Epitope Mature APC T cell epitopes are linear and directly derived from antigen sequence Binding is determined by amino acid side chains (R groups) and ‘encoded’ in single letter code 8
    9. 9. EpiVax HLA “Supertype” Coverage • EpiVax tests for binding potential to the most common HLA molecules within each of the “supertypes” shown to the left. • This allows us to provide results that are representative of >90% of human populations worldwide* without the necessity of testing each haplotype individually. * Southwood et. al., Several Common HLA-DR Types Share Largely Overlapping Peptide Binding Repertoires. 1998. Journal of Immunology. 9
    10. 10. EpiMatrix •The EpiMatrix algorithm scores all the 9-mers in a given sequence for binding affinity across a range of common HLA and reports both detailed and aggregated results. •In protein therapeutics and existing vaccines these tools can predict immunogenicity (efficacy). In the design of vaccines these tools allow for educated selection of pathogen genome sequences to make the most effective, efficient and safe vaccine 10
    11. 11. ClustiMer Easy • easy to deliver as peptides T cell epitopes are not randomly distributed but instead tend to cluster in specific regions. – These clusters can be very powerful, enabling significant immune responses to low scoring proteins. • ClustiMer recognizes T-cell epitope clusters as polypeptides predicted to bind to an unusually large number of HLA alleles. DRB1*0101 DRB1*0301 DRB1*0401 DRB1*0701 DRB1*0801 DRB1*1101 DRB1*1301 DRB1*1501 • T-cell epitope clusters make excellent vaccine candidates: – compact; relatively easy to deliver as peptides; highly reactive in-vivo 11
    12. 12. Conservatrix Finds Conserved 9-mers CTRPNNTRK CTRPNNTRK CTRPNNTRK CTRPNNTRK CTRPNNTRK CTRPNNTRK CTRPNNTRK Conserved epitope Identifying the most conserved 9-mers allows for protection against more strains with fewer epitopes 12
    13. 13. BlastiMer: Epitope Exclusion Foreign Self In all of our vaccines we eliminate cross-reactive epitopes 13
    14. 14. New iVAX feature: JanusMatrix Homology Analysis JanusMatrix is designed to predict the potential for cross reactivity between epitope clusters and the human genome, based on conservation of TCR-facing residues in their putative HLA ligands. TCR MHC 14
    15. 15. JanusMatrix Cross-reactivity Networks A different human protein A human protein Source Protein or large peptide Another human protein FLQDSNLYK (T cell epitope with same TCR face) VLQSSGLSYS ( T cell epitope) Source protein Predicted 9-mer epitope from a source protein 9-mer from human prevalent proteome, 100% TCR face identical to source epitope Human protein where cross-reactive epitopes are present
    16. 16. Example: HCV Vaccine 22/23 epitopes vs. human proteome HCV_G1_NS2_732 HCV_G1_1941 HCV_G1_DEXDC_1246 HCV_G1_1605 HCV_G1_NS2_748 HCV_G1_NS4B_1769 HCV_G1_2941 HCV_G1_2440 HCV_G1_2898 HCV_G1_NS4B_1725 HCV_G1_ENV_359 HCV_G1_2485 HCV_G1_NS4B_1876 HCV_G1_NS4B_1910 HCV_G1_ENV_255 HCV_G1_2879 HCV_G1_NS4B_1790 HCV_G1_NS4b_1798 HCV_G1_2913 HCV_G1_NS5A_1988 HCV_G1_2840 HCV_G1_NS2_909 All induced Teff response
    17. 17. Example: HCV Vaccine One “special” epitope: cross-reactive vs. human proteome vs. human microbiome 648590236 6 4 8 5 5HMPREF9551_01014 7349 HMPREF9534_04988 conserved 648583178 conserved hypothetical4 9 5 0 4 6453 HMPREF9547_04386 hypothetical p ro te in hypothetical 6 4 4_010100004179 6 4 7 9 4 1 7 6 7 p ro te4 6 2 4E 6 4 3 in[Escherichia coli conserved p HMPREF0485_04604CSAG_01006 6 -1 ] inner ro te in [Escherichia coli 1 9 MS 6 4 [Escherichia6 0 8 conserved membrane4 8 5 3 2 8 5coli 5 7 0 8 0 8 yeaL [Klebsiella 6 9 -1 ] MS HMPREF9540_01467 te MS HMPREF9535_02392 1 7 5 -1 ] 6 4 8 5sp. 2 2 66 5 1 5 5 2 2 9 0 7 8 1_1_55] hypothetical 4 5p ro 4 9in 9 6 3 4 1conservedconserved HMPREF9541_04708 p ro te [Escherichia sp. HMPREF9538_06054 in S hiD9_010100003853 hypothetical hypothetical 4_1_40B] conserved hypothetical4 8 5 4 2 6sp. 6 [Citrobacter 4 7 conserved inner te 8 HMPREF9543_00699 6 p ro te in p ro6 4in5 8 5 7 5 7 hypothetical p ro te in 30_2] membrane4 8 5 2 8 2 7 9 [Escherichia coli [Escherichia coli HMPREF9553_01980 p ro te in [Klebsiella sp. conserved HMPREF9536_02724 protein [Shigella MS ] hypothetical [Escherichia coliMS 9 2 -3 ] hypothetical4 8 5 5 8 9 4 4 1 1 5 -18MS 97 8 -1 ] 6 sp. D9] hypothetical 5 3 4 7 7 64 p ro te in p ro te in MS 1 1 6 -1 ] HMPREF9550_00991 in pHMPREF9548_02962 ro te [Escherichia coli [Escherichia coli [Escherichia coli conserved hypothetical 6 4 8 5 6 6 2 6 7 6MS52 0 0 -11 48 9970 ] MS 1 4 6 -1 ] hypothetical 8 4 -1 ]p ro te in MS HMPREF9530_03710 HMPREF9549_04771 p ro te in [Escherichia coli conserved 6 4 8 5 5 2 coli MS 1 8 2conserved -1 ] [Escherichia 1 5 4 hypothetical hypothetical HMPREF9531_05139 MS 1 8 7 -1 ] 64 p ro conserved p ro te in6 2 0 8 7 5 2te in ESAG_02468 [Escherichia coli hypothetical [Escherichia coli 6 4 8 2 2 8 4-1 ] MS p ro te in 5MS92 1conserved 1 8 5 -1 ] HMPREF9552_03152 [Escherichia coli hypothetical p ro te in MS 4 conserved 5 -1 ] [Escherichia sp. hypothetical p ro te in 3_2_53FAA] 647938633 HMPREF0485_01509 hypothetical p ro te in [Klebsiella sp. 1_1_55] 650540371 650565199 CCU_24070 ROI_08320 Predicted Predicted hydrolase integral (metallo-beta-lactamase membrane superfamily) p ro te in [Coprococcus [Roseburia sp. A RT55/1] intestinalis M 5 0 /1 ] 648582837 HMPREF9547_04045 general secretion pathway protein L [Escherichia coli MS 175-1] 644158927 646274622 HOLDEFILI_00670 644393128 BSIG_014661 8 8 2 647445068 65052 hypothetical HMPREF0539_2484 646897627 p e p tid yl-p ro lyl 6 4 4 3 5 5 ABC CUO_16121 0 9 AL1_16290 p ro te in6 5 1 5 5 4 8 7 1 6 5 0 5 5 9cis-tra n s 696 LRHM_1901 Na HMPREF0352_21497 0 CPA2 family HMPREF9439_02418 6443 transporter,2 8 4 hypothetical [Holdemania 644327138 643156423 monovalent /H a n tip 65 82 ABC superfamily 6 5 0 0 7FPR_256200 0p ro2 8in 7 6 5 4 isomerasete 0 HMPREF0530_2293 permease permease,o rte r 6505 HMPREF0530_0979 PRABACTJOHN_02052 6 8 3 5 7 Predicted 6 4 8 9 2filiformis VPI 5 (H [Lactobacillus HMPREF9475_01584 [Bacteroides sp. ATPro6 4in e --tRNA binding cation:proton 0 5 0 HMPREF9474_01708 YjgP/YjgQ p g lycin 9 2 6 3 4 4 te 3 [Alistipes shahii J1-31B-1, RO1_04000 ApbE protein putative efflux membrane HMPREF0189_02116DSM fa mily GG, hypothetical hypothetical Lparp8_010100001688 ) a n tip o rte r-2 rhamnosus 1_1_6] 8301] cassette ligase [Enterococcus WAL 1 hypothetical [Lactobacillus protein, MATE p permease 2 0 4 2 ] ATCC 53103] [Lactobacillus transporter, p ro te inro te in p ro te in [Lactobacillus [Parasutterella faecalis Glycyl-tRNA PC4.1] p ro te in paracasei ATCC fa mily [Faecalibacterium YjgP/YjgQ excrementihominis synthetase rhamnosus membrane ATCC paracasei beta [Clostridium [Clostridium 651517449 [Roseburia 2 5 3 0 2 ] [Parabacteroides p ra u [Burkholderiales LMS2-1] symbiosum p ro te2 5 b u n it in 3 0 2 ] su symbiosum sn itzii YIT 11859] HMPREF9412_4232 johnsonii DSMintestinalis 646219318 bacterium 3] (G lycine--tRNA WA L -1 4 1S L3/3] L -1 4 6 76 5 1 3 6 9 2 5 0 [Enterococcus 6 3 ] WA peptidase XB6B4] 18315] HMPREF0127_00159 faecium beta BSAG_02678 1_1_47] 6 5 1 3 5 6 8 7 2 ligase propeptide and 6 4 3 1 4 5 76 4 6 2 6 0 6 2 2 05 hypothetical conserved TX1330] subunit) HMPREF1017_00908 (GlyRS) 646007290 6 4 8 9 0YpeB domain 5767 646221716 PROVALCAL_01382 p ro te hypothetical [Lactobacillus in hypothetical OFBG_00434 BacD2_010100023154 p ro te HMPREF9010_03587in 2 4 7 4 6 BSCG_00263 Uncharacterized protein [Bacteroides sp. te in 648894538 6462 paracasei 647454257 AraJ p ro [Paenibacillus p ro te in putative phage transcriptional conserved membrane-associated 1_1_30] BSCG_03264 HMPREF0106_04079 [Bacteroides sp. CUY_0074 PKD [Bacteroides 8 7 0 0 :2 ] [O xalobacter 6 integrase sp. regulator HGF5] hypothetical 4 6 0 0 6 1 6 5 p ro0 0informigenes 6 5 te 6 3 5 0 9 transcriptional conservedD1 ] d o ma in p ro 6 5in 3 5 7 8 5 8 te 1 ovatus BacD2_010100017435 sp. [Bacteroides 6 4 6 0 0 6 sp. 6 4 4 0 9 8 7 4[Providencia 5 [Bacteroides 6 6 8 p ro te in HMPREF0864_04057 hypothetical regulator [Bacteroides 3_8_47FAA]PROPEN_03458 OXCC13] HMPREF1017_01886 6 4 7 8 7BacD2_010100020019 9 9 6 7 3_1_23] [Bacteroideshypothetical D2 ] sp. alcalifaciens hypothetical [Bacteroides sp. p ro te in ovatus SD CMC hypothetical p ro te in putative HMPREF0102_01841 Uncharacterized 30120] 2_2_4] DSM p ro te in [Bacteroides sp. D2 2 ] 3 f] p ro te in [Bacteroides sp. conserved regulatory membrane-associated [Enterobacteriaceae 2_2_4] [Bacteroides D2 ] p ro te in hypothetical protein [Proteus bacterium ovatus p ro te in [Bacteroides sp. penneri ATCC 9_2_54FAA] 651550285 3_8_47FAA] D2 ] [Bacteroides sp. 35198] HMPREF9538_04048 2_1_22] fimb ria l p ro te in [Klebsiella sp. MS 9 2 -3 ] [Escherichia coli 6 647453329 46225070 MS 1 9 8 -1 ] CUY_0572BSCG_03588 two response -co mp o n e n t 64 6 4 8 9 sensor 7 8 7 8 1 3 5 regulator system 0 5 3 7 5 HMPREF0102_00037 HMPREF9010_04162 receiver domain histidine two -co kinase/response mp o n e n t p ro 3 in two -co mp o n e n t 6 5 1te5 8 6 1 1 system system sensor 4 2 7 0 4 [Bacteroides regulator 6 4 3 1 1 9 5 7 3 HMPREF1017_02631 sensor 6474 [Bacteroides histidine BACDOR_00482 ovatus SD CMC histidinesp. CW3_1134 hypothetical kinase/response 2_2_4]4 6 2 1 6 3 f] kinase/response 8 6 7 2response Uncharacterized p ro te in regulator BSAG_02040 6 4 7regulator, 9 p ro te in [Bacteroides4 5 6 2 4[Bacteroides 0 0 4 5 4 5 6 4regulator 6 sp. h yb rid two CW1_1394-co mp o n e n t domain conserved in receiver ovatus BacD2_010100009346 2_1_22] system t (o n e -co mp o n e nsensorp ro te in response bacteria 3_8_47FAA] two -co mp o n e n t system) regulator histidine [Bacteroides [Bacteroides system sensor kinase/response [Bacteroides receiver domain sp. xylanisolvens dorei DSM histidine 6 4 8 5 8 9 9 9 16 4 8 5 9 7 1 4 3 3_1_23] 7 3 6 7 8 p ro te in 1regulator SD CC 1b] 65 3 17855] kinase/response HMPREF9551_00769 HMPREF9549_02210 [Bacteroides sp. [Bacteroides HMPREF0127_04532 regulator, conserved conserved 648538381 ovatus SD CC D1 ] hypothetical h yb rid hypothetical hypothetical 64 2a] p ro te in n e -co mp o n e n t7 4 5 6 1 2 3 HMPREF9548_01865 ('o p ro te in 6 4 8 5 2 9 4 0 36 4 8 5 8 2 9p ro te in hypothetical [Bacteroides sp.system') CW1_3815 03 648569110 [Escherichia coli 6 4 6 2 0 coli e fflu x HMPREF9536_03850 [Escherichia 7 5 2 5 te in p ro 1_1_30] HMPREF9547_04111 [Bacteroides sp. MS 1 9 6 -1 ] MS HMPREF9535_00693 651369358 ESAG_01252 transporter, hypothetical hypothetical 1 8 5 -1 ] [Escherichia coli D2 ] conserved HMPREF0127_00262 u te r o p ro te in MS 2 -1 ] p ro te in conserved 1 8hypothetical 648903495 hypothetical membrane 64 6 [Escherichia [Escherichiahypothetical 5 3 5 1 3 8 8 coli 8 5 6 4 9 9 coli 64 HMPREF9010_01082in 2 7 3 8 0 6 4 6 2 2 0 8 3 1 p ro te in p ro te6 2 HMPREF9530_02439 64 factor 648 MS 8 4 -1 ] MS 1 7 5 -1 ] p ro te in E 4_010100013599 5 6 2 0 6 7 o u[Bacteroides sp. te r BSCG_05865BSAG_04177 [Escherichia coli 6 conserved 4 8 5 2 5 2 1sp. HMPREF9550_04116 [Escherichia 2 lip o p ro te in , 647 hypothetical membrane4 5 3 2 7 1 te r HMPREF9552_05381 MS 7 8 -1 ] 1_1_30] ou hypothetical o 6 4 7 te r 3_2_53FAA] ro te in conserved CUY_5065u NodT family 4 4 2 6 6 0 p e fflu x p ro te in membrane membrane p ro te in conserved 4 8 5 3 2 1 sp. 6 CW3_4757 hypothetical [Bacteroides [Escherichia 2 2 O prM e fflu x x p ro te in fflu x p ro te in [Escherichia hypothetical coli HMPREF9540_00738 4 7in 5 6 e fflu ovatuse 85 transporter, 4 7 8 7 9SD 4 e fflu x 4_1_40B] 6 4p ro te 5 6 7 4 CC [Bacteroides sp. o p rM MS 6 4 8 5 ] 8p ro 6 in conserved 2 1 -1 8 0 6 te o p rM 2a] transporter, HMPREF9531_00538 [Escherichia coli HMPREF0102_01619 3_1_23] o u te r [Escherichiahypothetical coli [Bacteroides sp. HMPREF9553_04291 o u te r MS 1 8 7 -1 ] conserved membrane o u[Bacteroides sp. te r 4 MS61 8 5 5 2 4 4 7 D1 ] hypothetical 9 8 -1 ] p ro te in hypothetical factor 2_2_4] membrane membrane HMPREF9534_00084 p ro te in [Escherichia5colip ro2 in 6 4 8 7 7 6 4 te factor lip o p ro te e fflu7 5p ro te8 8 9 4 9 4 0 6 conserved 6 4 in0 0 x 4 8 4 in 6, [Escherichia coli HMPREF9541_04124 coli MS 1 [Escherichia NodT family HMPREF0106_04483 o p rM lip o p ro te in , hypothetical 1 5 -1 ] BacD2_010100024426 MS 2 0 0 -1 ] NodT MS [Bacteroidesu te r p ro te in conserved 4 5 -1 ] [Bacteroides sp. te rfamily ou o 6 4 5 3 4 5 9 6 8 4 6 2 6 7hypothetical 6 45 ovatus SD CMC 2_1_22] [Bacteroides [Escherichia coli4 membrane membrane 646895903 S hiD9_0101000091226 9 -1 ] p ro te in ESCG_00515 xylanisolvens 3 f] x p ro tee fflu x p ro te in MS 6 e fflu in LRHM_02324 3 9 2 7 7 4 1 hypothetical conserved SD CC 1b] [Escherichia coli Lparp8_010100008715 oprM precursor O prM conserved protein [Shigella hypothetical 1 1 6 -1 ] MS [Bacteroides sp. [Bacteroides sp. hypothetical Predicted sp. D9] p ro te in membrane D2 2 ] D2 ] p ro te in [Escherichia sp. [Lactobacillus p ro te in 1_1_43] 6443 6 6 4 4 [Lactobacillus9 1 4 4 1 651357900 47457612 rhamnosus 3 9 0 7 5 0 GG, HMPREF0539_0815 paracasei CW1_2658 HMPREF0539_0126 HMPREF1017_01928 ATCC 53103] 8 7 :2 ] permease0 0permease hypothetical efflux ABC 6 4 [Lactobacillus 4 [Lactobacillus3 2 8 0 2 3 p ro te in transporter, 648558895 HMPREF0530_1852 648581795 6 4 7 permease rhamnosus rhamnosus [Bacteroides 4 5 4 3 2 7 HMPREF9550_00942 LMS2-1] 6 4 6 0 0 6 1 2 6 CUY_0145 in4 7 8 8 2 1 8 0 HMPREF9547_03002 conserved p ro te 6 LMS2-1] ovatus sulfate ABC hypothetical BacD2_010100017240 sulfate ABC HMPREF0102_04029 647856891 efflux ABC 64 3_8_47FAA][Bacteroides transporter, 5 3 4 8 8 2 4 ABC transporter ovatus SD CC transporter, HMPREF0847_01428 6 4 6 2 3 1 3 7 0 p ro te in transporter, ABC transporter 6 4 8 5 3 6 3 5 4 6 4 8 5 3 6 9 2E 4_010100000800 7 permease permease [Lactobacillus permease 4 1 0 6 7 2a] permease 64854 GNAT family 6 4 7 4 permease su lfa HMPREF9548_00411 te /th io su lfa te 9 4 4 4 BSEG_03838 HMPREF9540_04974 protein CysT paracasei ATCC [Bacteroides sp. p ro 6 4in[Bacteroides sp. protein CysT HMPREF9531_02427 acetyltransferase CW3_3396 te 6 2 1 8 9 5 2 8 9 0 4 6 4 2 carbohydrate sulfate ABC sulfate ABC 5 5 1 coli 64 [Escherichiatransporter 651 663 D2 ] [Escherichia coli sulfate ABC [Streptococcus 0 0esterase family 2 5 3 0 2 ] efflux [Bacteroides 2_1_22] ABC BSAG_02315 650 5282 transporter,1 8 7 -1 ]su b u n it transporter, HMPREF9538_05427 MS HMPREF9010_02237 MS 1 7 5 -1 ] transporter, sp. 2_1_36FAA] transporter, SD 0 5 2 9 ABC transporter ovatus 5 4 CMC HMPREF9011_03782 te in 1 2 p ro permease 6 4 8 5 6 7 6 4 4 sulfate ABC permease [Escherichia sp. 6 5ABC transporter 1 647888687 6 4 7 9 3 7 permease7 8 6 9 3 2 permease 4_1_40B] 4 26 4 8 5 permease 3 f] hypothetical [Bacteroides proteinHMPREF9530_05088 CysT protein CysT HMPREF9445_01605 permease transporter, sulfate ABC HMPREF9541_05175 6 4 8 8 8 2 4 2 3 p ro teHMPREF0105_1086 HMPREF0485_00139 protein CysT p ro teefflux ABC [Bacteroides sp. in[Bacteroides sp. [Escherichia coli in d o re i [Escherichia coli 6 4 8 permease 526848 sulfate ABC sulfate ABC HMPREF9007_01172 5 _carbohydrate D16 4 8 8 9 4 4 0 2 [Bacteroides [Bacteroides sp. 3 6 /D4 ] 1_ MS 1 1 5 -1transporter,8protein CysT [Escherichia coli ] MS 1 2 -1 ] transporter, ] 3_1_23] 6 4 3 1 family HMPREF9536_01291 5 9 5 3 7 8 5 -1 ] permease hypothetical 3_1_40A]esterase2 2 4 7 0 transporter, transporter, MS 4 xylanisolvens HMPREF0106_03941 permease [Klebsiella 6 4 8 ABC transporter 1 2 p ro te in sulfate ABC sp. protein CysT permease CysTpermease p ro6 4in4 3 7 5 6 7 BACDOR_03380 te 7 SD CC 7 p 6 6485 6 4 6 2 2 11b] ro te in 9 MSHMPREF9549_00444 5 5 2 0 5 9 2 -3 ] permease CUU_2038 [Bacteroides sp. Predicted [Escherichiatransporter, sulfateHMPREF9534_02842 coli protein CysT [Bacteroides sp. [Klebsiella sp. ABC BSCG_003435 1 3 7 3 1 4 2 [Bacteroides 6 [Bacteroides sp. 6 2 1 -1 ] 3 0 3 8 5 8 1 0 3 1 64 GDSL-like 3_1_33FAA] xylanase/chitin MS 4 8 5 9permease 9transporter,sulfate ABC 1_1_55][Escherichia coli 1_1_14] ABC transporter YIT HMPREF0127_04005 ] clarus D2 2 HMPREF9551_02082 HMPREF9553_05328 p ro4 2in1 9 6 3 deacetylase protein CysT permeasetransporter, ] 6 4 te 7 MS 1 1 6 -1 permeasehypothetical 12056] sulfate ABC sulfate ABC [Escherichia coli6 4 6 2 0 9 5 8 permease 7 [Bacteroides 6 4 7 9 4[Bacteroides ROSEINA2194_03461 0528 protein CysT 7 8 5 4 6 5 2 64 [Bacteroides sp. p ro te in transporter, 8 4 -1 [Escherichia protein CysT vulgatus7PC510] 5 dorei MStransporter, ] 64 53 647 HMPREF0485_03374 0 8 7 2 1 0 1 6 0 DSM hypothetical coli ESAG_03285 HMPREF9543_04581 2_2_4][Bacteroides sp. 645280677 permease permease HMPREF0219_2871 1 7 8 5 5 ] HMPREF0220_2248 oxaloacetate p ro te in MS 1 8 5 susulfate ABC -1 ]lfa te su lfa te /th io[Escherichia coli B 2_010100009578 1_1_30] 6 protein CysT4 8 5 7CysT protein 2 0 3 9 ABC5superfamily 6 5 0 5 4 3 8 8 [Roseburia decarboxylase, ABC superfamily 648521295 transporter, transporter MS putative 2', HMPREF9535_03623 4 5 9 9 0 6 9 -1 ] [Escherichia coli [Escherichia coli 3 1 64 ENC_40020 ATP ATP inulinivorans beta subunit binding binding HMPREF9552_01462 ABC subunit CysT permease 646288901 3 '-cyclic sulfate 0 -1 ] MS ] 650590 cassette hypothetical2cassette 1 6 5 0 0 5 9 8 5 9 1 9 6 -1 MS 2 0PROVALCAL_01668 [Klebsiella sp.5 5 16841] 0 3 9 4 1 DSM 6 4 4 conservedtransporter, [Escherichiaprotein CysT sp. HMPREF0619_03414 nucleotide sulfate ABC ES1_16010 ATP transporter p ro te in 1_1_55] transporter HMPREF0864_00463 p ro te in COPCOM_03663 d o ma in 3_2_53FAA] [Escherichia coli conserved 2'-phosphodiesterase permease transporter, [Enterobacter synthase superfamily superfamily sulfate ABC 648600222 648543645 [Escherichia protein 4 5 8 8 5 coli 6 2 CysT MS 1 4 6 -1 ] hypothetical 64 hypothetical 6 4 7 8 7 6 4 0 1 [Bacteroides sp. permease subunit C. cloacae cloacae ro te in p ro te in p HMPREF9543_01698 transporter 6 4 5 3 4 4 3 0]7 HMPREF9549_05292 p ro te in MS 1 9 8 -1CSAG_02233 p ro teHMPREF0103_2298 in 2_1_7] [Escherichia coli protein NCTC 9394] [Eubacterium [Clostridium [Clostridium [Enterobacteriaceae cytochrome Ccytochrome C S hiD9_010100000825 7 8 -1 ] lfa te CysT su lfa te /th io su[Providencia [Parabacteroides MS 6 4 6 [Coprococcus 8472 conserved 648888683 siraeum 8 9difficile NAP08] difficile NAP07] biogenesis bacterium te /th io su lfa te su lfa biogenesis transporter sp. D13] hypothetical LRHM_2731 ATCC HMPREF0104_02520 6 5 1 comes alcalifaciens V10Sc8a] 5 5 0 6 1 2 p ro 8 6 4 8 5 7 3 2 6 0 in 9_2_54FAA] p ro te in 6 4 8 5 4 7 6te 1 transportersubunit CysT 27 transcriptional 7 5 8 ] p ro te in HMPREF9538_04375 5'-nucleotidase HMPREF9535_04845 DSM 30120] HMPREF9531_00663 [Escherichia coli su b u n it [Escherichia4coli 6462098 0 [Citrobacter sp. regulator [Bacteroides sp. family protein transporter, cytochromecytochrome C C [Shigella sp. D9] 30_2] MS 1 8 5 -1 ] ESAG_03538 MS 1 4 6 -1 ] 6 4 8 9 02_1_33B] 9135 [Lactobacillus small biogenesis biogenesis 6 4 7 4 5 5 2 1 3 [Bacteroides sp. fo rma te -d e p e n d e n t HMPREF9008_02106 rhamnosus GG, 3_1_19] conductance p ro 8 in 6 4 8 5 8 5 3 2 6 4te5 8 2 0 5 4 CUY_4106 nitrite reductase p ro te in 5'-nucleotidase mechanosensitive [Escherichia 6 4 4 3 9 1 1ATCC 53103] 06 HMPREF9553_01549 coli [Escherichia HMPREF9547_03262 coli conserved [Escherichia sp. 6 3 5 family protein 645346 HMPREF0539_0481 ion channel MS 7 8 -1 ] cytochromecytochrome C C hypothetical S3_2_53FAA] MS 4 5 -1 ] hiD9_010100012465 [Bacteroides sp. xre family MscS family biogenesis biogenesis p ro 6 4 6 2 2 2 2 7 5te in fo rma te -d e p e n d e n t 20_3] transcriptional p ro te in 6 4 p ro6 6 in5 2 4p ro8 9in 4 0 6 2 te 3 6 8 5 te 6 [Bacteroides 6 5 0 5 9 6 7 8 4 BSCG_00818 nitrite reductase [Klebsiella sp. regulator [Escherichia coli HMPREF9551_00418 [Escherichia coli ovatus CMC 6 4 5 3co mp le x 5 1 9 4 0 ESCG_00846 CK3_12330 peptidase9SD 2 65135 50 [LactobacillusMS 9 2 -3 ] conserved ] MS 1 7 5 -1 C 3 f] [Bacteroides sp. E 4_010100016373 MS 2 0 0 -1cytochrome ] Uncharacterized subunit NrfF HMPREF1017_03509 rhamnosus6 4 7 8 9 6 9 1 0 hypothetical biogenesis fo rma te -d e p esp. e n t0 7 2 6 Mo b A -re la te d 2_2_4] [Shigella 6n d D9] hypothetical 4 8 5 6p ro te 6 4 8 5 3 2 1 te3 LMS2-1] in p ro 9 in HMPREF0969_00842 nitrite reductase p ro te in p ro te in HMPREF9550_02773 [Escherichia[Escherichia coli sp. HMPREF9540_00809 conserved 645278865 [Clostridiales [Bacteroides 6 4 8 5 2 9 9co3 le x 1 mp cytochrome C 1_1_43] MS 1 9C -1 ] cytochrome 6 hypothetical B 2_010100000532 subunit NrfF sp. 6 5 0 0 1 0 9 4 5 7 9 3 8 7 7 4 SS3/4] ovatus HMPREF9536_04360 biogenesis biogenesis p ro te in 6 4 putative [Escherichia sp.ro te 5 3 HMPREF1007_03145 p6 3_8_47FAA] cytochrome4C 5 3 9 0 1 3 4 8 in7 3 8 2p ro te in HMPREF0485_01650 6 8 HMPREF9541_00303 [Bacteroides sp. endonuclease/exonuclease/phosphatase 4_1_40B] hypothetical 648908282 biogenesis [Escherichia coli HMPREF9548_02498 [Escherichia coli D2ro]te intranscription family protein HMPREF9008_01652 p0 MS 1 8 7 -1 ] MS p ro tecytochrome cytochrome C 1 1 5 -1 ] in regulatory C [Bacteroides sp. putative [Bacteroides sp. [Escherichiabiogenesis biogenesis coli protein opdE 62_1_7]5 4 6 4 4787 6 4 7 3 6 7 7 7 2 4_1_36] endonuclease/exonuclease/phosphatase p ro te in MS 8 4 -1 ] p ro te in [Klebsiella sp. EDWATA_00503 familyHMPREF0103_1383 protein [Escherichia coli 6 4 6 2 8 6 4 8 5 conserved [Escherichia coli 1 1 6 -1 ] 6 5 11_1_55]5 55190 th ia min [Bacteroides sp. MS HMPREF0619_01034 MS 1 8 2 -1 ] metabolism 4 5HMPREF9538_05669 20_3]hypothetical 6 351468 conserved hypothetical p ro te in associated E 4_010100013999 hypothetical [Bacteroides sp. p ro te in putative p ro te in p ro te in8 8 8 72_1_33B] 64 083 [Klebsiella sp. [Edwardsiella MFS-family [Parabacteroides HMPREF0104_00922 MS 9 2 -3 ] tarda ATCC tra n sp o rt 651557268 endonuclease/exonuclease/phosphatase sp. D13] 2 3 6 8 5 ] p ro 651554194 HMPREF9442_02006te in [Bacteroides sp. [Escherichia sp. HMPREF9439_01741 hypothetical 3_1_19] putative p ro te in 4_1_40B] membrane [Paraprevotella p ro te xylaniphila YIT in 648924658 648889363 1 [Parasutterella 1841] HMPREF0189_01578 HMPREF0104_03199 excrementihominis putative conserved YIT 11859] membrane 6 4 8 9hypothetical 10841 651549390 p ro te in p ro te in HMPREF9008_01966 64 [Burkholderiales HMPREF9538_031528 5 7 7 9 7 6 [Bacteroides sp. conserved r o dHMPREF9541_04458 bacterium 6 4 6 2 8 9 hypothetical 055 3_1_19] 1_1_47] HMPREF0619_03567 in7 7 2 5 1 647 sh a p e -d e te rmin in g r o d p ro te 8 646206839 sh a protein RodA p e -d e te rmin in g conserved HMPREF0103_3135 [Bacteroides sp. ESAG_00573 6 4 5 3 4 sp. 5 [Klebsiella7 5 0protein RodA conserved hypothetical 20_3] 648 5 S hiD9_0101000168415 5 6 6 8coli r o d [Escherichia MS 9 2 -3 ] p ro te in hypothetical sh a p ] cell HMPREF9534_04323 te rmin in g wall MS 1 1 6 -1e -d e 645282361 [Parabacteroides p ro te in 4853 sh a p e -d e te rmin in gr o d protein6RodA 1 2 7 4 B 2_010100018018 sp. D13] [Bacteroides sp. sh a p e rmin in g HMPREF9536_05721 6 4 8 [Shigella -d e te [Escherichia sp. hypothetical 2_1_33B] protein 5 4 1 7 5 2 648598735 ro 3_2_53FAA] d HMPREF9548_05239 RodA sp. D9] protein p ro te in HMPREF9549_03804 sh r o d6[Escherichia coli a p e -d e te rmin in g 48535600 rod [Bacteroides sp. sh a p e -d e te rminMSa6 9 -1 ]e te rmin in g RodA in g p e -d HMPREF9540_04218 protein 2 2 3 4 2_1_7] sh 64857 [Escherichia coli protein RodA r o dprotein RodA 6 4 8 5 6 5 3 6 2 MS HMPREF9535_03818 [Escherichia 2 1 e te rmin in g 6 5 5 1 2 coli [Escherichia coli 8 4 -1 ] -d 6 4 8 5 8 8 7 6 5 4 8sh a p e HMPREF9530_02805r o d MS 1 8 2 -1 ] HMPREF9531_04206 MS 648553070 HMPREF9553_04990 protein RodA r o1 8 5 -1e -d e te rmin in g sh a p ] d rod [Escherichia coli HMPREF9534_00707 rod protein 6 4sh a p e -d6 te rmin in g RodA 859393 e sh a p in g MS 1 g 8561740 conserved sh a p e -d e te rmin e -d e te rmin in1 5 -1 ] 6 4[Escherichia coli protein RodA HMPREF9551_04716 RodA HMPREF9550_03789 4 hypothetical 6 4 6 2 4 4 0 4 86 protein 9 7 0 [Escherichia coli 7 8 -1 ] protein RodA 8 5 2 4 MS rod HMPREF9552_05139 [Escherichia coli p ro te in CSAG_00429 coli sh a p e -d e te MS 2 1g ]r o d [Escherichia rmin in -1 648599190 r o 4 -1 ] 6 4 8 3 6 6 0 [Escherichia coli cell wall 0 -1 ] MS d 5protein 8 5sh a p e -d e te rmin in g MS 2 0 RodA HMPREF9549_04259 protein HMPREF9547_04869RodA MS 6 9 -1 ] sh a p e -d e te sh a p eg e te rmin in g rmin in -d [Escherichia coli conserved 350729 p ro7 9in9 2 8 86 4 5RodAMS 1 9 6 -1[Escherichia coli rod 6 4 te 3 protein ] 6 4 8 5 5 0hypothetical 252 E 4_010100010287 MS g coli [Citrobacter [Escherichia sh a p e -d e te rmin in1 8 7 -1 ] HMPREF0485_02161 sp. p ro te HMPREF9531_03236 in cell -1 ] 30_2]o d MS 1 9 8 wall protein RodA 646210728 r conserved [Escherichia coli sh a p e -d e te rmin in g [Escherichia coli sh a p e -d e te rmin in g ESAG_04421 MS hypothetical 1 8 5 -1 ] p ro te in MS 1 7 5 -1 ] conserved p ro te in protein RodA [Escherichia sp. hypothetical [Klebsiella sp. [Escherichia coli 4_1_40B] p ro te in MS 4 5 -1 ] 1_1_55] [Escherichia sp. 3_2_53FAA] 648520000 643923830 HMPREF9552_00166 HcinC1_010100012020 transporter, tetracycline major facilitator 643137611 resistance protein 6 4 3 9 2 8structuralfamily DESPIG_00391 977 Lparp8_010100014860 [Escherichia coli efflux A rabinose protein TetA MS 1 9 8 -1 ] 6 tetracycline [Helicobacter 4 3 1 5 8 8 9 9 6 4 8 5 7 9 5 1 4 permease resistance PRABACTJOHN_04529 [Desulfovibrio HMPREF9547_00720 cinaedi CCUG 64 002 structural 3 1 1 3A rabinose effluxtransporter, 18818] piger ATCC 6 4 3 9 1 CLONEX_00508 673 permease 0 9 8 ] facilitator protein9TetA 29 major Cbac1_010100030692 [Parabacteroides 3 0 8protein A rabinose efflux 2 1 6 4 4 0 family 6 4 6 6 4 6 79 5 [Lactobacillus tetracycline 4 4 2 8 8 2 5 9 PROPEN_00011 6 permease johnsonii DSM BSBG_04744 paracasei [Escherichia coli resistance ] HMPREF0497_13831 5 ] efflux tetracycline 83 8 7 0 0[Clostridium 1A rabinose MS 1 7 5 -1 ] :2 permease conserved 6 4 5 2 8 3 1structural6 9 9 7 4 2 7 resistance 4 7 6 44 2 2 6 4 8 5 4 5nexile DSM protein TetA 1 7 8 7 hypothetical [Proteus penneri B 2_010100021971 BSCG_05908 p ro te in HMPREF9543_03552 ] ATCC 35198] [Clostridiales p ro te in tetracycline transporter,9 1 3 [Bacteroides sp. tetracycline 0 643 32 sp. 1_7_47FAA] [Lactobacillus6 resistanceloniC5_010100009852 4 4 1 0 0 2 9 7 resistance 9_1_42FAA] major facilitator B buchneri COPCOM_00016 p ro te in family proteinte in ATCC p ro tetracycline 9 1 64 [Bacteroides 8 5 3 6 resistance3 A1 1 3 3 0 efflux 6 4 8[Escherichia coli1 5 7 7 ] rabinose 5 3 sp.[Bacteroides sp. B permease HMPREF9548_02020 bifN4_010100009360 2_1_7] MS 1 4 structural 6 -1 ] 2_2_4] tetracycline transporter, protein TetA [Coprococcus resistance ATCC comes major facilitator [Bifidobacterium 2 family protein n g u m lo nstructural 7 7 5 8 ] lo gum protein TetA [Escherichia coli CCUG [Bifidobacterium 52486] MS 1 8 2 -1 ] bifidum NCIMB 41171] 646266592 ESCG_01081 peptidase M48 [Escherichia sp. 1_1_43] 650020844 HMPREF1004_02309 peptidase M48, Ste24p [Ralstonia sp. 5_7_47FAA] 651352667 648564583 HMPREF0987_02151 HMPREF9530_02026 hypothetical bacterial p ro te in extracellular [Lachnospiraceaelu te -b in d in g so 651551067 bacterium protein, family HMPREF9538_04831 9_1_43BFAA]5 [Escherichia ABC coli MS 21-1] transporter, 648534633 HMPREF9540_03250 peptidase, M48 fa mily [Escherichia coli MS 1 1 5 -1 ] su b stra te -b in d in g 647941285 protein, family HMPREF0485_04125 5 [Klebsiella sp. fimb ria l MS 9 2 -3 ] mo rp h o lo g y [Klebsiella sp. 1_1_55] 647367697 650273052 EDWATA_00428 HMPREF9406_4121 zinc transporter PRD domain ZupT p ro te in [Edwardsiella [Clostridium sp. tarda ATCC 6 5 1 5 1 6 3 5 9HGF2] 8 4 3 0 6 6513 236 HMPREF9412_4202 HMPREF9477_01558 8 5 ] 647369369 EDWATA_02100 putative ABC transporter, permease p ro te in [Edwardsiella tarda ATCC 23685] transporter, hypothetical major facilitator p ro te in HCV_G1_NS2_794 family protein [Lachnospiraceae [Paenibacillus bacterium sp. HGF5] 2_1_46FAA] 651543799 HMPREF9446_01544 polysaccharide biosynthesis p ro te in [Bacteroides fluxus YIT 12057] 650055436 HMPREF0177_01823 643911022 ribosomal L5P B bifN4_010100007817 family protein 50S ribosomal 650528031 [Bifidobacterium 643913229 protein L5 BIL_04240 LSU sp. B loniC5_010100009395 ribosomal [Bifidobacterium 12_1_47BFAA] 50S ribosomal bifidum NCIMB protein L5P protein L5 [Bifidobacterium 41171] [Bifidobacterium lo n g u m lo n g u m lo n g u m lo n g u m F8] CCUG 52486] 650558585 FPR_13180 Type IV 6 5 0 5 4 5secretory 0 1 8 7 4 6 6 835 65 pathway, EC1_18790 VirD4 HMPREF9436_01203 8 6 4 5 5 6501 651382993 Response components /TraD TraG HMPREF9436_00192 HMPREF9477_00264 [FaecalibacteriumproteinTraG /TraD regulators family hypothetical consisting ra ua itzii p of [Faecalibacterium protein sn family p ro te in S cf. CheY -like L3/3] prausnitzii 0 0 3 6 0 9 [Faecalibacterium 646 [Lachnospiraceae receiver domain KLE1255]cf. prausnitzii BacD2_010100004621 bacterium KLE1255] 6 42_1_46FAA] and a 7881619 putative win g e HMPREF0102_03473 d -h e lix 6 5 1 3 5 transmembrane 1291 DNA DNA -b in d in g HMPREF0987_00810 tio n mo b ilisa 643114071 651516134 646222494 topoisomerase I d o ma in HMPREF9412_2818 CLONEX_01578[Eubacterium hypothetical p ro te in BSCG_01035 6 4 5 26 4 7 4 8 9 3 2 0 hypothetical sp. 74522 [Bacteroides p ro 5 1 3 [Bacteroides sp. 6 5 0 3 te in 1 1 650274500 HD domain conserved B fra3_010100003045 2_1_22] 0 4 4 1 6478765 HMPREF0281_00213 p ro te in 6 2 4cylindroides[LachnospiraceaeD2 9 3 HMPREF9406_1660 64 HMPREF9443_01091 ] p ro te in hypothetical T2 - 8 7 ] putative 6bacterium 3 4621352 HMPREF0103_2485 glutamate 6 4 3 9 1 4 8 3 2 2 5 2 3 4 3 [Clostridium 6MBAG_00279 46 hypothetical hypothetical [Paenibacillus esterase sp o rt BSBG_01832 ABC transporter p ro te in p ro te in tra n FSAG_01313 9_1_43BFAA] nexile DSM two -co p ro te in sp. HGF5]Cbac1_010100006573 mp o n e n t 6 5 1 5 1 6 8 0 sp. [Bacteroides 3 [Bacteroidesin d in g conserved permease A TP -b 1 7 conserved 87 [Clostridium sp. HMPREF9412_0790 a u xin e fflu x ] system 6 5[Phascolarctobacterium 2_2_4] 1 3 4 sp.6YIT212067]5 8hypothetical 4 1 65 6 7 3 4 6 fragilis 3_1_12] in 4 8 5 3 5 7 2 9 ro te p hypothetical 4 [Bacteroides sp.pyridine HGF2] carrier transcriptional 650560145 HMPREF0992_00365 in p ro te 2_1_33B] HMPREF9540_04347 [Corynebacterium BSIG_00304n u cle o tid e -d isu lfid e ro te in [Clostridialesp regulator hypothetical FPR_30670 644157867 PRD domain 5 2 1 3 7 3 [Bacteroides sp. 650 ammoniagenes1_7_47FAA] [Fusobacterium conserved oxidoreductase [Coprobacillus sp. conserved 644182000 9_1_42FAA] p ro te in AL1_09950 DSM 20306] RUMHYD_035417 8 6 p ro te in hypothetical sp.42_1_31] 6 3sp. 5 1 3 D7] [Paenibacillus hypothetical putative efflux [Lachnospiraceae CLOSTASPAR_06618 [Escherichia coli hypothetical p ro 4 in COLSTE_011506 sp. HGF5] 6 4 6 0 4 0 1 0 bacterium 6 4 7 4te8 7 8 1 ro 0 in protein, MATE CAAX9aminop 6 5te 1 8 7 8 4 0 MS 1 1 5 -1p ro te in ] 648 25517 CUW_0440 Response 6 [Bacteroides sp. 643148124 ChatD1_010100029034 4 8 9 2 7 3 9 9 6_1_63FAA] 1_1_6] te rminHMPREF9436_01578 aTIGR00043 l [Alistipes shahii family [Blautia regulators PROVALCAL_03811 hydrogenotrophica -d e p e n d e n t DNA -b in d in6 0 5 1 5 5 8 5 0 0 HMPREF0326_00253 b in d in g -p ro te in HMPREF0326_02114 8 g 6 651570 protease[Faecalibacterium family. conserved WAL 8301] ir o n - su lfu r p ro in consisting of a hypothetical CDP-alcohol teHMPREF9442_03238 [Clostridiumrahypothetical tra HMPREF9413_1869 p u sn itzii DSM 10507]n sp o rt [Turicibacter clu ste r-b in d in g p ro CheY -like 65 177 p ro te in systems phosphatidyltransferase inner putative hypothetical 0 5 2 1asparagiformeS L3/3] te in 7513 AL1_07600 p ro te in TIGR00043 6 4[Providencia 8926749 receiver domain6 4family 1 1 7sanguinis protein membrane aliphatic p ro te in DSM 15981] HMPREF0493_1380 [Desulfovibrio PC909] HMPREF0326_01475 and o Predicted [Faecalibacterium alcalifaciens co mpa n e n t [Desulfovibrio sulfonates [Paraprevotella Na /H 64 the rh o mb o id 30120] win g e d -h e lix sp. 3_1_syn3]n3 9 1 3 9 1 5 hydrolase ofsp. 3_1_syn3] prausnitzii DSM fa mily [Clostridium cf. tra sp o rt xylaniphila Cbac1_010100001995 YIT p ro te in DNA -b in d in g a n tip o rte r permease metallo-beta-lactamase KLE1255] hathewayi DSM 11841] [Desulfovibrio superfamily d o ma in9 ][Lactobacillus diguanylate 1347 protein SsuC cyclase/phosphodiesterase [Alistipes shahii sp. 3_1_syn3] [Collinsella amylolyticus [Paenibacillus [Clostridiales stercoris DSMDSM 11664] sp. HGF7] WAL 8301] sp. 1_7_47FAA] 13279] 647459210 CW1_1639 RNA polymerase sig ma -7 0 fa cto r [Bacteroides 646275464 ovatus SD CC BSIG_02296 2a] RNA polymerase ECF-type sigma factor [Bacteroides 7 6 8 6 5 1 3 7 3 sp. 1_1_6] HMPREF0127_04622 hypothetical 648892415 p ro te in HMPREF0106_01939 [Bacteroides sp. RNA polymerase 1_1_30] ECF-type sigma factor 648881902 [Bacteroides sp. HMPREF9007_00606 D2 2 ] RNA polymerase ECF-type sigma factor [Bacteroides sp. 1_1_14] 644227085 651350965 EUBHAL_01530 HMPREF0987_00485 electron hypothetical tra n sp6 4 3 1 1 5 4 1 0 o rt p ro te 4 6 1 0 8 0 4 5 6 in CLONEX_02919 co mp le x, [Lachnospiraceae HMPREF0494_0359 electron RnfABCDGE bacterium conserved tra n type, D subunit sp o rt 9_1_43BFAA] hypothetical co [Eubacteriummp le x, p ro te in 651349687 6 5 1 3 6 hallii6DSM33353] 8 2 7 15 1 3 RnfABCDGE 5340 [Lactobacillus HMPREF0992_01853 HMPREF0991_01506 HMPREF1025_02141type, D subunit antri DSM hypothetical [Clostridium hypothetical hypothetical 16041] p ro nexile DSM te in p ro te in p ro te in [Lachnospiraceae 1 76 7 ] 8 [Lachnospiraceae 5 1 3 8 3 4 5 6 [Lachnospiraceae 6 5 1 3 2 8HMPREF9477_00720 7 7 4 bacterium bacterium 6 5 0 0 4 7 8 0 5 bacterium 6_1_63FAA] HMPREF0990_00997 2_1_58FAA] 2 7 6 6 3_1_46FAA] HMPREF1026_00500 6 4 4 1 0hypothetical hypothetical p ro te COPCOM_02487 in Na-translocating p ro te in [Lachnospiraceae electron NA DH-q u in o n e [Lachnospiraceaebacterium reductase3 3 0 7 0 6 tra n sp o rt 6 5 1 bacterium 2_1_46FAA] co mp le x, [Lachnospiraceae HMPREF0988_00134 1_1_57FAA] RnfABCDGE bacterium sp. hypothetical type, D subunit 8_1_57FAA] te in p ro 644180003 [Coprococcus [Lachnospiraceae CLOSTASPAR_04620ATCC comes bacterium Response 2 7 7 5 8 ] 1_4_56FAA] 643112755 regulators 6 4 6 2 CLONEX_00261 650556253 41223 consisting of a MBAG_01060 FP2_19490 Response CheY -like 65 conserved 6 9 Response 0 5 6 2 6regulators receiver domain hypothetical MHY_04970 regulators consisting of a and a Response pa consisting of ro te in CheY -like win g 6 5 0 5e lix8 5 7regulators e d -h 5 9 CheY[Coprobacillus -like receiver domain FPR_27460 DNA -b in d in g consisting of6a 0 1 8 8 1 5 8 5 sp. 1 8 4 2 6 1 receiver domain D7] and a Response 6 4 3 d o ma in CheY -like g e HMPREF9436_01896 andEUBIFOR_01499 d -h e lix win regulators a [Clostridium receiver domain putative KDP win g e DNA -b5 8d in g consistingd -h e lix 6 4 1 in 2 6 asparagiforme of a Response 4 operon 3 DNA -b in d in gand a d o ma in regulators DSM CheY -like win g eHOLDEFILI_00005 15981] d -htranscriptional e lix d o ma in consisting[Clostridium of a receiver domain Response g 6 5 0 0 2 7 0DNA -b nexile DSM 72 [Faecalibacterium in d inregulatory and a 6 CheY -like regulators d o ma inprotein HMPREF1011_00516 domain7 8 7KdpE 6 4 3 1g e d -h6 lix4 4 0 9 6 9 9 01 p ra 8 e receiver consisting ]of a win 4 7 7u sn itzii PROPEN_01698 [Megamonas response PROVALCAL_03473 and [Faecalibacterium a CheY -like DNA -b L 2 -in ] Response in d 6 g cf. regulatorhypermegale Responsein win g e d -h e lixprausnitzii d o ma receiver domain A RT1 2 /1 receiver regulators ] KLE1255] regulators DNA [Faecalibacterium -b in d in g d o ma in -co nconsisting of aand a ta g consistingsn itzii in ino ma in of a d p ra u win ro te CheY -like g e d -h e lix CheYpL3/3]in[Eubacterium S-like receiver domain-b in d in g DNA 6 4 8 5 3 0 2 9 2 receiver domain [Anaerostipes biforme 6 5 1 3 3 4 6 9 0 DSM HMPREF9536_04739 sp. 3_2_56FAA]and a d o ma in and a ] HMPREF0991_00859 win g3 9 8 9e lix e d -h[Holdemania hypothetical win g e d -hhypothetical e lix filiformis VPI DNA -b in d in g p ro te in DNA -b in d in p ro te in g J1-31B-1, DSM domain [Proteus [Escherichia coli d o ma in 648584330 [Lachnospiraceae 1 2 0 4 2 ] penneri ATCC MS 8 4 -1 ] HMPREF9553_00553 [Providencia bacterium 9 8 ] 351 alcalifaciens hypothetical 2_1_58FAA] DSM 30120] p ro te in [Escherichia coli MS 2 0 0 -1 ] Induced Treg response 64621849 646228165 6 4 6 2 1 3 4 5 9 BSEG_00669 6 4 3 1 2 3 0 4 8 BSBG_01768 ABC transporter BACDOR_03958 ABC transporter A TP -b in d in g ABC-type A TP -b in d in g p ro te in mu ltid r u g p ro te in [Bacteroides tra n sp o rt [Bacteroides sp. d o re i system, ATPase 644392137 9_1_42FAA] 5 _ 1 _ 3 6 /D4 ] and permease4 3 2 8HMPREF0539_1504 64 538 HMPREF0530_2361ro n e ctin fib components fib b 6 4 3 9 1 7 8[Bacteroides ro n e ctin in d in g p ro te in 6 8 9 7 1 0 7 88 64 A [Lactobacillus Cbac1_010100021787 DSMd in g p ro te in dorei b in LRHM_1393 A ] hypothetical 1 7 8 5 5[Lactobacillusrhamnosus n e ctin -b in d in g fib ro paracasei ATCC LMS2-1] p ro te in p ro te in 25302] [Clostridiales [Lactobacillus9 2 6 4 0 7 643 sp. 1_7_47FAA] rhamnosus GG, Lparp8_010100002015 ATCC 53103] -b in d in g fib ro n e ctin 647369118 EDWATA_01849 th re o n in e -p h o sp h a te decarboxylase [Edwardsiella tarda ATCC 23685] p ro te in 647369715 [Lactobacillus EDWATA_02446 646897150 integral paracasei LRHM_1436 0 :2 ] membrane 8 7 0 ATP-dependent p ro te in exonuclease [Edwardsiella subunit A 6500632 tarda ATCC7 1 [Lactobacillus 2 64439218 HMPREF0864_03827 23685] HMPREF0539_1549 rhamnosus GG, UDP-N-acetylglucosamine ATP-dependent ATCC 53103] d ip h o sp h o ryla se /g lu co sa min edeoxyribonuclease, -1 -p h o sp h a te N-acetyltransferase subunit A [Enterobacteriaceae [Lactobacillus bacterium 6 4 3 9 2 6 3 5 9 rhamnosus Lparp8_010100001765 6 4 7 8 5 59_2_54FAA] 848 LMS2-1] ATP-dependent HMPREF0847_00391 exoDNAse hypothetical 6 4 4 3 2 8 4 8 6 (exonuclease V) p ro teHMPREF0530_2309 in beta subunit [Streptococcus ATP-dependent [Lactobacillus sp. 2_1_36FAA] deoxyribonuclease, paracasei 643155446 subunit A 7 0 0 :2 ] 8 PRABACTJOHN_01075 [Lactobacillus RagB, SusD and paracasei ATCC hypothetical 25302] 650592773 proteins. CL3_32810 [Parabacteroides TRAP 643138141 johnsonii5DSM5 9 1 64 282 651312886 650533738 transporter, DESPIG_00923 6 5 0 0 1 8 8 0 4 6 4 8 8 8 9 72_010100019185 5 B 21 8 3 1 5 ] 648576449 CLS_16740 subunit HMPREF0240_01840 DctM HMPREF0104_03558 hypothetical HMPREF9541_02931 Glycogen HMPREF1004_00251 shikimate TRAP [Clostridiales RagB/SusD p ro te in debranching twin -a rg in in e conserved transporter,S M4/1] kinase sp. enzyme d o ma in -co n ta in in g translocation [Bacteroides sp. hypothetical DctM subunit p ro te in [Desulfovibrio pathway signal [Clostridium sp. 7 9 3 8 7 5 9 64 2_1_7] 646289358 p ro te in [Clostridium cf. D5 ] piger ATCC 7 6 9[Ralstonia sp. [Bacteroides sp. 6505 16 HMPREF0485_01635 6 4 8 9 0 8 6HMPREF0619_03868 01 [Escherichia coli saccharolyticum 3_1_19] 2 9 0CK1_14550 98] HMPREF9008_03392 5_7_47FAA] 6 5 1 5 5 1 9 1 1 transcription conserved MS 1 1 6 -1 ] K 10] hypothetical regulator RagB/SusD hypothetical HMPREF9538_05675 protein, MDR hypothetical p ro te in p ro te in LysR substrate [Ruminococcus e fflu x p u mp p ro te in [Parabacteroides b in d in g d o ma in sp. SR1/5] cluster [Bacteroides sp. sp. D13] p ro te in 20_3] [Klebsiella sp. [Klebsiella sp. 1_1_55] MS 9 2 -3 ] 643916645 Cbac1_010100015578 PTS system, IIB co mp o n e n t [Clostridiales sp. 1_7_47FAA] 650063005 HMPREF0864_03563 PTS protein [Enterobacteriaceae bacterium 9_2_54FAA] 646223825 BSCG_02356 conserved hypothetical 6 4 8 8 9 2 2 5 1 1 3 6 9 6 5 0 p ro te in 65 HMPREF0106_01775 [Bacteroides sp. HMPREF0127_00550 647450958 2_2_4] conserved 7 4 4 2 1 4 4 64 hypothetical CUY_4616 hypothetical p ro te in CW3_0806 conserved te in p ro conserved hypothetical [Bacteroides sp. [Bacteroides sp. hypothetical 1_1_30] p ro te inD2 2 ] 648588549 p ro te in [Bacteroides 645345425 HMPREF9553_04774 [Bacteroides S hiD9_010100006402 itrite ovatus SD CMC n xylanisolvens 648566899 3 f] n itra te /n itrite SD CC 1b]transporter transporter HMPREF9530_04343 64857038 [Escherichia0coli [Shigella sp. D9] 2 0 0 -1 ] n itrite HMPREF9535_01964 MS transporter n itrite 6 4 6 2 0 8transporter 6 4 8 5 4 1 1 1 6 194 [Escherichia coli ESAG_01916 HMPREF9548_04602 [Escherichia coli4 6 4 8 5 4 7 2 4 MS 2 1 -1 ] n itrite e xtruMS n7 8 -1 ] n itrite 5 5 8 0 5 6 sio 6 HMPREF9531_002264 8 transporter p ro te in 1 n itrite HMPREF9550_00103 [Escherichia coli [Escherichia sp. n itrite transporter 8 2 -16 4 8 5 7 8 7 8 7 ] 3_2_53FAA]1 6 4 8 MS 1HMPREF9541_05269 30 78 64859590 [Escherichia5coli 3transporter HMPREF9536_04825 n coli HMPREF9549_00967 5 -1 ] [Escherichia itrite MS 4 n itrite6 4 8 5 8 0 5 -1 ] MS 1 8 7 6 5 n itrite transporter transporter [Escherichia coli HMPREF9547_01772 transporter3 5 0 0 6 3 645 n itrite [Escherichia coli MS 1 1 6 -1 ] [Escherichia coli E 4_010100006994 648545466 MS 8 4 -1 ] transporter MS 1 8 5 -1 ] n itra te HMPREF9543_03519 /n itrite 6 4 8 5 2 3 6 6 6 [Escherichia coli n itrite transporter 6 4 8 5HMPREF9552_03834 [Escherichia 9 3 2 8 9 MS 1 7 5 -1 ] sp. transporter n itrite HMPREF9551_04069 [Escherichia coli7 5 6 6 4 8 5 5 6 4_1_40B] n itrite transporter MS 1 4 6 -1 ] HMPREF9534_04394 [Escherichia coli transporter n itrite MS [Escherichia coli 1 9 8 -1 ] transporter MS 1 9 6 -1 ] [Escherichia coli MS 6 9 -1 ] 648579916 648568952 HMPREF9547_01122 HMPREF9535_00535 P px/G ppA 6 4 8 5 3 6 5 0 1 P px/G ppA phosphatase HMPREF9540_05121 phosphatase family protein P px/G ppA family protein 6 4 8 5[Escherichia coli 94024 phosphatase [Escherichia coli 6 4 5 3 4 8 4 1 4 HMPREF9551_048045 -1family protein2 6 8 2 6 2 ] 646 S ] MS 1 7 MS 7 8 -1hiD9_010100021332 P px/G ppA guanosine ESCG_02094 [Escherichia1 6 4 8 5 3 8 0 0 coli phosphatase a n o sin eMS 1 1 5 -1sp h a te ,3 '-d ip h o sp h a te g u HMPREF9548_01485 -5 '-trip h o ] pentaphosphate family protein 9 6 3 4 7 pyrophosphatase 6phosphohydrolase ppA 485 P px/G 648544703 [Escherichia coli HMPREF9549_01414 [Escherichia sp. [Shigella sp. D9] HMPREF9543_02756 phosphatase MS 1 9P px/G ppA 6 -1 ] 1_1_43] family protein 0P px/G ppA 64621124 phosphatase ESAG_04930 [Escherichia coli phosphatase 64 574481 family8protein MS 1 h a te],34 8 5protein g u a n o sin e -5 '-trip h o sp8 2 -1 familyip5 4 7 0 6 a te 6 '-d h o sp h HMPREF9541_00961 [Escherichia pyrophosphatase coli HMPREF9534_02343 [Escherichia coli P 1 8 5 -1 ]6 MSpx/G ppA 4 8 5 2 4 5 4 3 P px/G ppA ] [Escherichia sp. 1 4 6 -1 MS 6 4 8 5 8 5 0 7 7 HMPREF9552_04712 phosphatase 3_2_53FAA] HMPREF9553_01300 family protein2px/G ppA phosphatase 1P 8 6 4family protein P px/G ppA 6 4 5 3 5 3phosphatase8 5 6 0 3 0 8 [Escherichia coli E 4_010100022326 HMPREF9550_02355 [Escherichia coli phosphatase 1 1 6 family protein MS -1 ] guanosine MS 6 9 6 4 family6protein 8527 80 [Escherichia P px/G ppA -1 ] 648 3 coli 41 pentaphosphate 5 6 7phosphatase [Escherichia coli HMPREF9536_02124 MS 1 8 -1 ] HMPREF9530_04785 phosphohydrolase 9 family protein MS 2 0 6 4 8 5 5 0 4 5 3 P px/G ppA 0 -1 ] HMPREF9531_03437 [Escherichia coli [Escherichia P px/G ppA sp. phosphatase P px/G ppA phosphatase 1 8 7 -1 ] 4_1_40B] MS family protein family protein phosphatase [Escherichia coli [Escherichia coli family protein MS 8 4 -1 ] [Escherichia coli MS 2 1 -1 ] MS 4 5 -1 ] 650087089 HMPREF9464_00302 polar amino acid ABC transporter [Sutterella wadsworthensis 3_1_45B] 650060234 HMPREF0864_00824 650560164 sulfatase FPR_30930 [Enterobacteriaceae Restriction 650008741 endonuclease S bacterium 9_2_54FAA] HMPREF1007_00969 subunits hypothetical 643147537 [Faecalibacterium p ro PROVALCAL_03223 te in 6 5 0 3 5 3 9 5 4 p ra u sn itzii [Bacteroides sp. Predicted HMPREF9444_01560 S L3/3] membrane 4_1_36] conserved 644156633 p ro te in 646272574 d RUMHYD_02306 o ma in p ro te in [Providencia BSHG_03947 [Succinatimonas Pyruvate:ferredoxin alcalifaciens conserved hippei YIT oxidoreductase DSM 30120] hypothetical 1 2 0 66 5 1 3 6 3 2 3 3 6] and related p ro te in 2 -o xo a cid :fe rre d o xin HMPREF1018_02096 [Bacteroides sp. oxidoreductases, hypothetical 3_2_5] gamma subunit p ro te in [Blautia [Bacteroides sp. hydrogenotrophica 644286383 2_1_56FAA] 647893735 DSM 10507] HMPREF0496_2581 HMPREF9024_01405 conserved transcription 6 5 0 1 9 8 2 6 2 hypothetical accessory p HMPREF0623_0974ro te in p ro te in YhgF like[Lactobacillus [Pediococcus p ro te in brevis acidilactici 7_4] gravesensis [Pediococcus acidilactici DSM ATCC 27305] 20284] 646240811 MBAG_00648 predicted p ro te in [Coprobacillus sp. D7] 650020839 HMPREF1004_023041 5 4 6 2 5 6 65 HMPREF9538_00017 ATP-dependent Clp protease, negative A TP -b in d in g regulator of 644294383 648531805 643098504 subunit ClpC genetic HMPREF0348_1204 6 8 8 8 6 1 648 4 648 HMPREF9540_00421 5 3 4 8 3 sp. competence [Ralstonia 6 4 7 9 8 6 5 0 4 6 4 7 9 8 8 3 6 5 BACEGG_03273 650041378 possible HMPREF9521_00708 ATPase family HMPREF9540_03452 Mg 5_7_47FAA] ClpC/MecB HMPREF0873_00628 HMPREF0874_00617 -d e p e n d e n t HMPREF1016_00978 histidine kinase p ro te in ATPase family histidine kinase [Klebsiella sp. me th ylma lo n yl-Co A DNase me th ylma lo n yl-Co A Mg p ro te in [Enterococcus 8 6 1 1 [Escherichia 6 5 0 5 3 7 5 2 3 [Bacteroides -d e p e n d e n t [Enterococcus 65058 6 4 6 2 6 6 5 9 8 coli MS 9 2 -3 ] epimerase epimerase DNase faecalis MS 1 1 5 -1 ] [Escherichia coli CC1_22940 faecalis ENT_26790 ESCG_01087 [Veillonella sp. [Veillonella sp. eggerthii DSM 644177756 644209508 [Bacteroides TX0104] MS 1 1 5 -1 ] Uncharacterized TX2134] Signal 20697] ATPase with 6_1_27] CLOSTASPAR_02371 3_1_44] CLOSTMETH_03465 eggerthii conserved transduction TrpR-related chaperone 644270805 643139396 Predicted Zn 1_2_48FAA] protein (some activity p ro te in 4 3 9 1 7 0 9 4 ROSEINA2194_02303DESPIG_02179histidine kinase 6 peptidase 643124383 members 6 4 6 2 4 7 1 8 4 [Enterococcus YerC/YecD [Escherichia sp. Serine/threonine Cbac1_010100017829 O uter [Clostridium ANHYDRO_00256 4 6 2 6 5 8 8 9 contain a von CSAG_03511 6 sp. 7L76] 1_1_43] [Clostridium protein kinase TrpR like 5 0 5 9 8 5 9 1 membrane 6 methylpentosum Cna protein HWAG_01253 Willebrand Sua5/YciO /YrdC/YwlC asparagiforme ro te inCK3_33190 648927142 [Roseburia p , p ro te in R2, DSM 5476] B -type domain. conserved factor type A family 650088265 HMPREF0326_01859 protein inulinivorans DSM 15981] YerC/YecD TrpR-related 6 5 1 3 3 7 4 7 1[Anaerococcus hypothetical (vWA) domain) 651517389 HMPREF9464_01439 (O mpH-like). RNA [Citrobacter sp. DSM 16841] [Clostridialesp ro te in [Desulfovibrio HMPREF9412_5440 HMPREF0994_00057 hydrogenalis p ro te in [Coprococcus flp p ilu s mo d ifica tio n 30_2] sp. 1_7_47FAA] piger 5 1 6 7 8 0 0 6 6 ATCC YerC/YecD hypothetical DSM 7454] [Helicobacter transcriptional catus G D/7] assembly 646035840 enzyme, HMPREF9418_0568 29098] [Clostridiales p ro te in 6 4 4 0 winghamensis 98715 regulator, LuxR p ro te in ChatD1_010100007516 646261147 MiaB -family transcriptional sp. SS3/4] 651342555 650319738 [Lachnospiraceae fa mily PROPEN_03428 ATCC BAA-430] 6 5 0 0[Sutterella 61657 hypothetical OFBG_00955 [Desulfovibrio regulator A nr [Paenibacillus HMPREF0994_05103 bacterium HMPREF9420_0951 translation wadsworthensis p ro te in FA D/FMN-containing sp. 3_1_syn3] HMPREF0864_02232 4 6 3 0 4 9 9 9[Neisseria 6 hypothetical conserved 3_1_57FAA_CT1] sp. HGF5] factor SUA5 3_1_45B] alpha amylase 6 [Clostridium dehydrogenase HMPREF0026_024424 7ATCC4 7 macacae 4 4 9 7 p ro te in hypothetical [Proteus penneri [Enterobacteriaceae hathewayi DSM [O xalobacter 33926] p rimo so ma l CUW_1263 DNA [Lachnospiraceae p ro te in ATCC 35198] bacterium topoisomerase 13479] formigenes protein N 646224828 bacterium [Prevotella 6 5 0 3 5 1 5 2 7 9_2_54FAA] 643139916 6500 947 IV, A subunit 6 4 8 5 8 7 4 5 9 BSCG_03346 OXCC13] [Acinetobacter 3_1_57FAA_CT1]salivae1 2HMPREF9443_01307 DSM DESPIG_02699 HMPREF9553_03683 HMPREF0983_01497 junii SH205] [Turicibacter conserved 1 5 6 0 6 ] L-asparaginase 650352274 GTP sanguinis hypothetical hypothetical 644316707 putative 648547530 [Phascolarctobacterium HMPREF9443_02055 cyclohydrolase aspartate PC909] p ro te in p ro te in HMPREF0545_1139 HMPREF9531_00512 sp. YIT 12067] CRISPR-associated subunit MoaA ribonuclease HI aminotransferase [Escherichia6coli3 6 0 0 9 46 4 4sp. 6 8 4 8 [Bacteroides 2 2 iron chelate 51 protein, Csn1 [Lactobacillus [Desulfovibrio0 1 [Erysipelotrichaceae MSHMPREF1017_04090 2 0 0 -1 ] 2_2_4] EUBHAL_01293 uptake ABC 6 4 6 2 0 7 5 0 0 6 5 ATCC fa mily piger1 3 6 7 7 bacterium sp. salivarius HMPREF1025_01578 transporter, ESAG_01227 hypothetical Phosphate [Phascolarctobacterium 9 5 5 1 64625 29098] HO66, ATCC 3_1_53] sta rva tio n -in d u cib le FeCT 6 4 8 5 8 8 0 9 4 ABC family, hypothetical iron sp. YIT 12067] OFAG_01507 p ro te in 648926945 HMPREF9553_04319 5 4 3 2 8 6 11741] permease 650 643147789 [Bacteroidesprotein PhoH, p ro te in transporter chaperone SurA HMPREF0326_01668 predicted iron p ro te inchelate ENC_32680 ovatus [Lachnospiraceae PROVALCAL_03476 permease [O xalobacter 5 0 0 1 3 4 8 9 6 4 -(cytid in e uptake ABC ABC-type Fe3 ATPase [Escherichia coli bacterium Predicted [Escherichia sp. 644097630 formigenes 3_8_47FAA] 5 '- d ip h o sp h o ) - 2 - C- me th yl- D- e r yth r ito l HMPREF0983_02062 [Eubacterium MS transporter, -sid e ro p h o re 4 5 -1 ] 3_2_53FAA] 64414387 650087745 3_1_46FAA] 6 membrane PROPEN_02340 HOxBLS] putative kinase FeCT 5 9 7 1 1 6 tra n sp o rt hallii DSM 3353] BACCOPRO_00205 HMPREF9464_00950 6 4 8 family, p6 4te8 5 7 5 2 4 ro 7 in AAA domain [Desulfovibrio 6 6 diarrheal toxin 6475132 permease crp /Fn r fa mily HMPREF9549_02183 system, Uncharacterized HMPREF0847_02056 [Providencia (Cdc48 6 4 6[Erysipelotrichaceae 243637 648590015 sp. 3_1_syn3] HMPREF0493_1501 conserved p ro te in Transcriptional iron chelate permease alcalifaciensd in gsubfamily). A TP -b in HMPREF9551_00793 CSAG_00020 bacterium sp. hypothetical [Escherichia coli mp o n e n t regulator uptake ABCco p ro te in DSM 30120] [Proteus penneri transporter predicted 3_1_53] iron chelate p ro te in6 4 3 1[Bacteroides MS 2 0 0 -1 ] [Enterobacter 16437 transporter, [Sutterella ComA ATCC 35198] p ro te0 3 2 0 8 4 1 uptake ABC 6 4 6 8 9 6 2 8 3 CLONEX_03946 [Lactobacillus 6 5 in wadsworthensis FeCT 6 4 8cloacae8 family,4 9 7 cloacae coprophilus [Streptococcus 5 6transporter, 650045118 [Citrobacter sp. LRHM_0608 amylolyticus DSM 18228] HMPREF9420_2035 3_1_45B] ABC-type permeaseNCTC 9394] sp. 2_1_36FAA] HMPREF9530_02421 651572855 HMPREF1023_00790 30_2] 648552428 FeCT family, putative te /su lfo n a te /b ica rb o n a te DSM 11664] conserved n itra p ro te inchelate HMPREF9413_5814 iron luxR family HMPREF9534_00065 permease multidrug ABC hypothetical tra n sp o rt [Escherichia coli arylsulfatase Bacterial iron chelate uptake ABC te in p ro transporter 650073316 system, ATPase 6 5 0 5 3 9 6 6p ro te in [Paenibacillus 5 regulatory uptakeMS 1 8 5 -1 ] ABCtransporter, [Escherichia coli A TP -b in d in g [Prevotella HMPREF9488_01292 co mp o n e n t5 1 6 0 1 3 CCU_15010 651 transporter,FeCT family, 9 6 -1 ] 6 5 sp. HGF7] MS 1 co mp o n e n t 6 4 3 9 2 5 7 p ro te in 71 ABC transporter salivae DSM 1 3 8 3 6 2 2 [Clostridium hypothetical HMPREF9412_5193 FeCT family,permease 650062 HMPREF9477_00885 4 7 8 8 5 5 7 1 [Lactobacillus9 2 9 nexile DSM Hp u lM9 _ 0 1[Eggerthella 2 5 0 1 0 0 0 0 9 4 sp. ] 6 [Coprobacillus p ro te in 1 5 6 0 6hypothetical 1_3_56FAA] permease p ro te in HMPREF0864_03487 transcriptional thymidylate rhamnosus GG, HMPREF0101_02623 sp. 29_1] 1787] [Coprococcus [Escherichia coli regulator, 6 4 4 1 6 0 0 1 9 AraC p ro te in yrdC p ro te in ATCC 53103] kinase conserved 6 045277 sp. A RT55/1] 5 0[Lachnospiraceae MS fa mily [Escherichia coli 2 1 -1 ] d o ma in -co n taHOLDEFILI_01763 in in g [Helicobacter hypothetical HMPREF1023_00949 [Paenibacillus Predicted MS 6 9 -1 ] p ro te in bacterium pullorum MIT p ro te in luxR family 646269 sp. HGF5] [Enterobacteriaceae transcriptional 2_1_46FAA] [Bacteroides 5 1 2 98-5489] sp. Bacterial BSHG_00932 regulators bacterium 2_1_16] regulatory conserved 9_2_54FAA] [Holdemania 6 5 1 3 1 2 4 6 3 ro te in 647371096 650021631 p hypothetical 644160607 filiformis VPI 643 HMPREF0240_01414 sp. 6 5 1 3 6 1 8 7p ro te in EDWATA_03829 0 [Eggerthella HMPREF1004_03094 9 2 2 0 8 9 HOLDEFILI_02351 J1-31B-1, DSM HcinC1_010100003150 AMP-dependent dipeptide ABC HMPREF1018_00744 sp. 1_3_56FAA] [Bacteroides 6 5 0 5 3 8 3 3 0 Na -d rive n 1 2 0 4 2 ] DNA hypothetical synthetase and transporter, polymerase III hypothetical 3_2_5] CC1_32230mu ltid ru g e fflu x p ro te in ligase A TP -b in d in g subunit alpha p ro te in p u mp ATPase, P-type 51 [Helicobacter [Clostridium6sp. 5 6 7 8 7 1 [Bacteroides sp. protein DppD [Ralstonia sp. 647513128 (transporting), [Holdemania HMPREF9413_2216 646262918 D5 ] [Edwardsiella 5_7_47FAA]cinaedi CCUG 2_1_56FAA] filiformis VPI HMPREF0493_1391 HAD B eta-eliminating 2 0 6 65 tarda ATCC0 5 6 5 8 5 0 6 4 4 1 0 2 1 8 8 1 8 ]4 3 1 4 0 2 0 0 HRAG_00601 cytidylate superfamily, J1-31B-1, DSM COPCOM_02000 6 5 0 5 6 9 0 2 2 6 5 1 5 1 6 7 9 6 lyase DESPIG_02983 thymidylate 23685] ROI_15910 6 12042] 647442302 643145692 kinase 4 4 2 8 4 1 0 3 6 4 4 1 5 9 7 9 9 IC subfamily [Paenibacillus Transcriptional hypothetical tRNA(Ile)-lysidine kinase 64 8474 HMPREF9412_0783 HOLDEFILI_01543RO1_11940 HMPREF0496_0349 [Coprococcus CW3_4406 PROVALCAL_013694 1 5[Lactobacillus sp. hypothetical B eta-eliminating HGF7] p ro te in [Helicobacter synthetase, regulators HOLDEFILI_00216 648927374 Transcriptional adenylylsulfate hypotheticalD/7] conserved amylolyticus catus G bilis ATCC p ro te in N-te rmin a l [Coprococcus [Desulfovibrio Predicted 11664] lyase 650571179 p ro te in reductase d o ma in p ro te in HMPREF0326_02088 regulator DSM comes ATCC piger ATCC [Roseburia [Paenibacillus domain/tRNA (Ile)-lysidine RO1_37110 43879] type VI 6 4 3 1 4 0 2 1 5 [Providenciatranscriptional [Lactobacillus [Holdemania [Bacteroides 27758] 6 5 VPI intestinalis synthetase, tRNA(Ile)-lysidine 0 9 8 ] 29 DESPIG_02998 sp. HGF5] 6 filiformis 0 0 8 7 8 3 9 6 4 6xylanisolvens 6 0 0 6 6 224770 474 brevis secretion alcalifaciens regulators HMPREF9464_01029 XB6B4] C-te rmin a l His Kinase A synthetase, BSCG_03288CW1_4644 gravesensis J1-31B-1, DSM p ro te in , DSM 30120] [Holdemania SD CC 1b] hypothetical d o ma in (phosphoacceptor) 6 5 0 5 2 8 9 3 1 VPI filiformis conserved conserved N-te rmin a l 12042] ATCC 27305] VC_A0114 p ro te in [Roseburia d d J1-31B-1, DSM domain/tRNA (Ile)-lysidine fa milyo ma in ./Histid in e BIL_14850 hypotheticalo ma in p ro te in hypothetical 0 4 2 ] [Sutterella intestinalis kinase-, DNA 12 p ro te in [Bacteroides synthetase, [Desulfovibrio p ro te in wadsworthensis M 5 0 /1 ] gyrase B-, and ovatus SD 6 4[Bacteroides 7 8 8 0 0 0 8 CC 6 0 0 6 6 4 66 4 sp. C-te rmin a l sp. 3_1_syn3] 643095437 3_1_45B] HSP90-like [Bifidobacterium 2a] d o ma in HMPREF0102_01882 2_2_4] 6 4 8 9BacD2_010100019909 05787 BACEGG_00204 lo n g u m lo n g u m ATPase. hypothetical conserved [Roseburia HMPREF9010_03609 4 - a min o - 4 - d e o xy- L - a r a b in o se F8] [Desulfovibrio te intestinalis hypothetical 6 4 8 8 9 0 7 5 7 conserved 6p ro4 5in 5 1 7 6 5 0 0 4 2 2 2 0 transferase and 47 5 651542515 piger ATCC [Bacteroides sp. XB6B4] hypothetical CUY_1326 p ro te in HMPREF1016_01813 related HMPREF0106_00282 6441 29098] HMPREF9446_00260 6 5 7 6 4 D2 ] [Bacteroides sp. p ro 9 in glycosyltransferases hypothetical 6 4 6 2 1te2 7 4 conserved d oBACCELL_00927 -ma n n o se -p ro te in lich yl-p h o sp h a te d o lich yl-p h o sp h a te -ma n n o se -p ro te in 651541957 2_1_22] of p ro te in [Bacteroides sp. 4 - a min o - 4 - d e mannosyltransferase PMT family BSAG_02634 hypothetical mannosyltransferaseo xy- L - a r a b in o se HMPREF9445_03033 3_1_23] [Bacteroides [Bacteroides sp.conserved p ro te in transferase [Bacteroides te -ma n n o se -p ro te in [Bacteroides d o lichand h o sp h a eggerthii DSM yl-p eggerthii D2 2 ] hypothetical [Bacteroides fluxus YIT related mannosyltransferase 1_2_48FAA] 20697] p ro te in ovatus SD CMC 1 2 0 glycosyltransferases [Bacteroides 57] [Bacteroides sp. 3 f] of PMT family clarus YIT D1 ] [Bacteroides 12056] cellulosilyticus 650583643 DSM 14838] RTO_16000 TrpR-related 644156939 p ro te in RUMHYD_02612 651366495 YerC/YecD TrpR-related HMPREF1025_00393 [Ruminococcus p ro te in TrpR protein u e s L 2 -1 4 ] to rq YerC/YecD YerC/YecD 6 5 1 3 4 6 8 3 3 651337064 HMPREF0993_02418 [Blautia [Lachnospiraceae HMPREF0991_03195 hypothetical hydrogenotrophica 1 0 8 651328 6 4 4 2bacterium 70816 hypothetical p ro te in DSM 10507] HMPREF0990_00344 3_1_46FAA] 6513 ROSEINA2194_02314 3 3 1 4 4 p ro te in [Lachnospiraceae HMPREF0988_02534 6 7 7 4 2 6 5 0 0 4 7 3 7 1[Lachnospiraceae protein 6 5 0 5 7 0 3 1 8 TrpR TrpR-related 650 bacterium5 HMPREF1026_00070 bacterium YerC/YecD RO1_26580 p ro te in hypothetical ROI_38070 5_1_57FAA] TrpR-related[Lachnospiraceae YerC/YecD p ro te in TrpR-related TrpR/YerC/YecD 2_1_58FAA] 0 1 3 5 8 6441 bacterium p ro te in p ro te in [Roseburia [Lachnospiraceaep ro te in COPCOM_01077 6 1312216 6 5 1 3 4 3 2 YerC/YecD 1_1_57FAA] inulinivorans5bacterium YerC/YecD [Lachnospiraceae6 4 TrpR-related 650538389 bacterium sp. [Roseburia p ro te in DSM HMPREF0240_01166 CC1_32850 16841] 1_4_56FAA][Roseburia HMPREF0994_05806 4 hypothetical 3 1 1 2 5 1 Trp R-like 8_1_57FAA] 6intestinalis 5 YerC/YecD TrpR-related 6 intestinalis CLONEX_00021 p ro te in XB6B4] p ro te in 5 0 5 7 7 5 6 8 [Coprococcus M 5 0 /1 ] CK1_22230 p ro te in 0 5 8 0 5 7 2 TrpR-related [Lachnospiraceae [Clostridium sp. 65 comes ATCC YerC/YecD TrpR-related bacterium p ro te in D5 ] CK5_17800 27758] [Coprococcus p ro te in 3_1_57FAA_CT1] TrpR-related YerC/YecD catus G D/7] YerC/YecD [Clostridium p ro te in [Ruminococcus YerC/YecD nexile DSM sp. SR1/5] 1787] [Ruminococcus o b e u m A 2 -1 6 2 ]
    18. 18. EpiAssembler Produces Immunogenic Consensus Sequences STRAIN 01 STRAIN 02 STRAIN 03 STRAIN 04 STRAIN 05 STRAIN 06 STRAIN 07 STRAIN 08 STRAIN 09 STRAIN 10 STRAIN 11 STRAIN 12 STRAIN 13 STRAIN 14 STRAIN 15 STRAIN 16 STRAIN 17 STRAIN 18 STRAIN 19 STRAIN 20 Q Q Q Q Q Q Q Q Q Q Q Q Q Q Q Q X Q Q Q Q X A X A X A X A X A A A A A A X A X A A A S S S S S S S S S S S S S S S S S S S S S W W W W W W W W W W W W W W W W W W W W P P P X P P P X P P P X P X P P X P X P K K K K K K K K K x R K K K K K K K K K K V V X V V X V V X V V V V X V V V X V V E E E E E E E E E E E E E E E E E E E E Q X Q Q Q Q Q Q Q Q Q Q Q Q X Q Q Q Q Q F F F F F F F F F F F F F F F F F F F F W W W W W W W W W W W W W W W W W W W W A A A A A A A A A A A A A A X A A A A A K K K K K X K K K K K X K K K K K K K X H H H H H H H H H H H H H H H H H H H H X M M M M M M M M M M M M M M M M M M M W W W W W W X W W W W W W W W W W W W W N N N N N N N N N N N N N N N N N N N N X F F F F F F F F F F F F F F F F X F F I I I X I I I I X I I I I I I I I I I I S S S S S S S S S X S S S S S X S S S S X G G X G G G G X G G G G X G G G G X G I I I I I I I I I I I I I I I I I I I I Q Q Q Q Q Q Q Q X Q Q Q Q Q Q Q Q Q Q Q Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y Y L L X L L X L L X L L X L L L L X L L L Y L F W A K H M W N F M W N F I S G I Q W P K V E Q F W A W P K V E Q N F I S G I Q Q A S W P K V E Q F W A K H M W N F I S G I Q Y L
    19. 19. Epitope Cluster Score Junctional Cluster Score 20 10 0 -10 Peptides in Default order in construct HP_IIB 50 40 HP4117 HP4061 HP4181 HP4111 HP4018 HP4070 HP4060 HP4157 HP4065 HP4001 HP4193 HP4034 HP4068 HP4168 HP4160 HP4175 HP4127 HP4126 HP4007 HP4154 HP4164 HP4119 HP4100 HP4120 HP4179 30 EpiMatrix Cluster Score 50 HP4117 HP4179 HP4007 HP4111 HP4018 HP4070 HP4034 HP4193 HP4065 HP4181 HP4157 HP4060 HP4068 HP4164 HP4160 HP4175 HP4127 HP4120 HP4126 HP4154 HP4168 HP4119 HP4100 HP4001 HP4061 EpiMatrix Cluster Score VaccineCAD Epitope Cluster Score Junctional Cluster Score 40 30 20 10 0 -10 Peptides in Optimized order in construct HP_IIB VaccineCAD Eliminates Introduced Junctional Epitopes 19
    20. 20. Output: Multi-Epitope Gene Design Intended Protein Product: Many epitopes strung together in a “String-of-Beads” DNA insert DNA Vector Protein product (folded) 20
    21. 21. Multiple Delivery Platforms Possible DNA – chain of epitopes, or peptide in liposomes ICS-optimized whole proteins ICS-optimized proteins in VLP
    22. 22. iVAX: Case Studies 22
    23. 23. Case Study: Smallpox Vaccine VennVax smallpox vaccinia Immunogenic Epitopes Vaccine Shared Immunogenic Epitopes
    24. 24. ‹#›
    25. 25. Case Study: VennVax Immunization in HLA DR3 Transgenic Mice Immunizations Days 0, 14, 28, 42 Immunogenicity Day 56 1. epitope DNA vaccine prime (IM) 2. epitope peptide boost (IN) • Challenge done at 10X LD50 for Smallpox • Related publication: http://www.ncbi.nlm.nih.gov/pubmed/21055490 Moise L et al. Vaccine. 2011;29:501-11 Challenge Day 65
    26. 26. VennVax protects against lethal vaccinia challenge 100% survival of Vaccinated mice vs. 17% of placebo 100 100% 90 Percent Survival 80 Placebo 70 Vaccinated 60 50 40 30 20 10 DNA DNA boost boost Challenge 17% 0 00 100 20 5 60 10 40 15 Day Post Immunization 20 80 25 Moise L et al. Vaccine. 2011;29:501-11
    27. 27. VennVax Protection: Morbidity Percent of Pre-Challenge Weight 115 VennVax 110 Placebo 105 100 95 90 85 80 75 70 65 0 5 10 Days Post-Infection 15 20
    28. 28. Case Study: VennVax Protection: Antibody Titers VennVax conferred protection without any B cell epitopes Moise et al. Vaccine. 2011; 29:501-11
    29. 29. Other Vaccine Design Case Studies • 5 prototype vaccines using iVAX with 3 validated in animal models (VennVax, TulyVax, and H.pylori) • Validation of remaining prototype vaccines (FluVax) currently in progress 29
    30. 30. Immunogenicity of H7N9 HA predicted to be LOW Previous vaccines poorly immunogenic - 80 - - 70 - - 60 - - 50 - - 40 - HA A/California/07/2009 (H1N1) Tetanus Toxin - 30 - Influenza-HA HA A/Victoria/361/2011 (H3N2) - 20 - - 10 - - 00 - - -10 - - -20 - IgG FC Region - -30 - Fibrinogen-Alpha - -40 - - -50 - - -60 - - -70 - - -80 - Thrombopoietin Human EPO EBV-BKRF3 HA A/Texas/50/2012 (H3N2) Albumin Follitropin-Beta H7 HA Immunogenic Potential Random Expectation HA A/chicken/Italy/13474/1999 (H7N1) . . . . . . . . . -6.23 HA A/Shanghai/1/2013 (H7N9) . . . . . . . .. . . . . . . . -8.11 HA A/mallard/Netherlands/09/2005 (H7N7) . . . . . . -8.63 gB-2 (EPX Score: -24.56) HA A/mallard/Netherlands/12/2000 (H7N3) .. . . . . .-9.91
    31. 31. H7N9: Prediction of lowimmunogenicity IAV T Cell Epitope Responses by Category Average Immune Response N=4 Subjects EpiVax predictions H1N1 (2009) H7N9 (2013) T Cell Epitope Content1 HIGH VERY LOW T Cell Epitope Cross Reactivity2 MEDIUM VERY LOW Cross-Reactive Antibody Immunity3 LOW LOW Predictive protective effect of vaccination in standard influenza vaccine HIGH LOW In Novartis Phase I trial of H7N9 cell culture vaccine showed only 6% of 400 healthy volunteers achieved a protective response when given two doses of the 15ug un-adjuvanted vaccine
    32. 32. IVAX makes vaccines: More Effective: IVAX can be used to design more targeted, immunogenic vaccines or predict the immunogenicity of existing vaccines Safer: Epitope-driven DNA or Sub-unit vaccines mitigate the risks associated with live, attenuated or VLPs without sacrificing efficacy. Less antigenic load. Faster: After H7N9 genome was released EpiVax personnel were able to complete a vaccine sequence in less than 48 hours – approx 20 hours total labor. 32
    33. 33. Accessing the Tools Contact Jason Del Pozzo: bda@epivax.com PreDeFT: Fee for service in silico immunogenicity analysis. Performed on a protein by protein basis. Pricing based on length of sequence(s). Limited ISPRI Website: Limited access to EpiVax’ Interactive Protein Screening and Reengineering Interface. Available for set numbers of proteins. Unlimited ISPRI Website: Unlimited access to EpiVax’ Interactive Protein Screening and Reengineering Interface. Available in three year lease periods. Fee for Service: HLA Binding Assays, HLA Transgenic Mice, ELISpot Assays. Confidential 33

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