This document discusses the analysis of protein BAB71591 which is deregulated during Hepatitis C infection. The analysis involved: 1) Retrieving the protein sequence from a database and predicting its unknown function using bioinformatics tools. It was predicted to localize to the cytoplasm and function as a methyltransferase. 2) Building a homology model based on a phospholipase template to predict the protein structure. The model showed alpha helices and beta sheets. 3) Correlating the putative functions to Hepatitis C, finding they may be downregulated during infection through methylation or inflammatory responses.

1. In silico analysis of BAB71591 :a protein of unknown function deregulated during Hepatitis C in Homo sapiens PRESENTED BY: AFSANA M.Sc. Bioinformatics (IVth SEM) SESSION-2008-2009

2. HEPATITIS C AND HEPATITIS C VIRUS(HCV) Infectious disease affecting liver, caused by HCV . Acute hepatitis C-Persisting Less than 6 months. Chronic hepatitis C-Persisting More than 6 months. HCV - small , enveloped, single-stranded, positive sense RNA virus. Transmission through blood transfusions that are not screened for HCV infection, reuse of inadequately sterilized needles.

3. OBJECTIVES STEP 1 :To retrieve deregulated gene of unknown function. STEP 2: To predict function of the protein encoded by the same deregulated gene using different soft wares, tools and databases available in the public domain. STEP 3: To determine putative structure of the same protein. STEP 4: To find a correlation of the same hypothetical protein in the light of Hepatitis C disease.

4. STRATEGY FOR RETRIEVING GENE/PROTIEN STANFORD MICROARRAY DATABASE PICK GENE OF UNKNOWN FUNCTION FROM MICROARRAY EXPERIMENTS IN HEPATITIS C(HUMAN) CHECK WHETHER IT IS AN EST OR NOT IN dbEST YES NO BLASTX OF THE SEQUENCE RETRIEVED FROM dbEST RETRIEVED UNAMED PROTEIN FROM BLASTX RESULTS General information about retrieved EST EST name:z076d04 GenBank Acc.-AA158258 EST length -625 bp

5. STRATEGY FOLLOWED FOR FUNCTION PREDICTION UNNAMED QUERY PROTEIN NCBI BLAST P(NR) CDD INTERPRO DATABASE HYDROPATHY PLOT PFAM DATABASE FUNCTION OF THE UNNAMED PROTEIN PREDICTED PSORT

6. To retrieve general information of retrieved protein BAB71591 from NCBI Url : http://www.ncbi.nlm.nih.gov / Comments: ACCESSION NUMBER : BAB71591 ORGANISM : Homo sapiens PROTEIN LENGTH : 165 amino acid GENE SIZE : 1796 base pair GENE IDENTIFICATION NUMBER : AK057830

7. To retrieve related proteins to query protein in different organisms using BLASTP (database :nr) Url: http:// blast.ncbi.nlm.nih.gov/Blast.cgi Comments: The first related protein is SET-domain containing protein functions in signaling processes and related to methyltransferases.

8. To find localization of the hypothetical protein. Url: http:// wolfpsort.org / Comments: Localization was found in cytoplasm.

9. To find particular family to which hypothetical protein may belong to. Url : http:// pfam.sanger.ac.uk / Comments : 3 families were identified as: Family PFAM-B 83975 having domain Nol1_Nop2_Fmu Family –PFAM-B-197450 having domain SLA_LP_auto_ag Family PFAM_B 5481 having domains jMjC , ARID , PLU-1

10. WORK SUMMARY Tool/software/server used Comment/Conclusion BLASTP (database: nr) SET-domain containing protein-methyltansferase. CDD database No domain identified * Interpro database No significant result * Psort tool Cytoplasm Hydropathy plot Pfam database String tool Cytoplasmic localization * Three families were identified No interaction * Conclusion for primary sequence analysis: Protein is Cytoplasmic and may play function as methyltansferase. * - Indicates data not shown for particular results.

11. STRATEGY FOLLOWED FOR HOMOLOGY MODELING Selection of template Alignment of target and template Model generation Validation of model Template selected : crystal structure of human cytosolic phospholipase A2 PDB ID:1CJY Seq identity: 33% w.r.t BAB71591 Length of template : 749 residues Length of BAB71591:165 aa

12. To predict the model for protein BAB71591 using modeller Url: http:// www.salilab.org/modeller/download_installation.html Comments : Predicted model shows 3 helices and 2 beta sheets and coils. Predicted model for BAB71591

13. To validate the model for BAB71591 using PROCHEK Url : http://nihserver.mbi.ucla.edu/SAVES / Comments: Core region value-74.6% Disallloweds-0% Bad contacts - 0

14. To identify putative pockets in predicted model using ligsite server Url: http://gopubmed2.biotec.tu-dresden.de/cgi-bin/index.php Comments: Three pockets were identified. PKT373 PKT74 PKT48 POCKETS

15. CORRELATION OF PUTATIVE FUNCTION WITH HEPATITIS C PHASE I : PRIMARY SEQUENCE ANALYSIS Protein is cytoplasmic and can perform function as methyltransferase. PHASE II: STRUCTURE ANALYSIS Protein is cytoplasmic and can function as human phospholipase A2 enzyme. CONCLUSION: Protein can perform either of these functions. If methylation-methylation is down regulated in Hepatitis C. If Phospholipase responsible for inflammatory response it is also down regulated in Hepatitis C.