Meta-genomics is the application of modern genomics techniques to the study of communities of microbial organisms directly in their natural environments, bypassing the need for isolation and lab cultivation of individual species”
Protein engineering is the process of developing useful or valuable proteins. It is a young discipline, with much research taking place into the understanding of protein folding and recognition for protein design principles
Meta-genomics is the application of modern genomics techniques to the study of communities of microbial organisms directly in their natural environments, bypassing the need for isolation and lab cultivation of individual species”
Protein engineering is the process of developing useful or valuable proteins. It is a young discipline, with much research taking place into the understanding of protein folding and recognition for protein design principles
We're creating awesome websites and applications using MODX, but being a developer usually brings one downside: we're bad at empathising with our end-users. This talk is about optimising MODX for your end-users. I'll be talking about must-have extra's, easy-to-implement form customisation, brilliant TV's, which you should always use, and of course, a healthy portion of SEO.
Improved coverage of the proteome using gel eluted liquidExpedeon
It has long been understood that sample fractionation is critically important to generating quality, comprehensive proteomics data. In spite of the continual improvements in speed and sensitivity of mass spectrometers, these instruments are still unable to adequately overcome the enormous challenge
of most biological samples without multiple dimensions of separation prior to mass analysis.
Oncology: Spatial Localization of Ras proteinsNachiket Vartak
This is a presentation of work done at the MPI Dortmund from 2008-2013 on the mechanism through with localization of the Ras protein in generated in cells. It presents the inhibiton Palmostatin-B, which inhibits this mechanism, leading to reveral of oncogenic signaling and cancerous phenotypes.
Application of proteomics for identification of abiotic stress tolerance in c...Vivek Zinzala
It is the study of “Proteome”.
The word "proteome" is a blend of "protein" and "genome”.
Large scale study of Proteins.
Particularly their structures and functions.
Study of full set of proteins in a cell type or tissue, and changes during various conditions
Crimson Publishers- CPG Methylation in G-Quadruplex and IMotif DNA StructuresCrimsonPublishers-SBB
Abberant hypomethylation in DNA regions with noncanonical folding potential (ncDNA motifs) is believed to predetermine tumor development - presumably, by facilitating G-quadruplex (G4) and/or i-motif (IM) formation via altering nucleosome positioning (stable G4s induce subsequent genomic rearrangements). We questioned whether CpG methylation per se affects the dsDNA-ncDNA equilibrium. Thermodynamic studies of genomic and model oligonucleotides with methylated CpG sites at different positions are reported. The genomic oligonucleotides analyzed in this work are DNA fragments with reportedly different methylation statuses in colorectal cancer and normal cells. Free energies of duplex, ncDNA formation from single strands were calculated based on melting curve analyses. Polyethylenglycole was used to imitate crowding effect. Our results suggest that CpG methylation may alter the energetic barrier for dsDNA-IM transitions.
The IOSR Journal of Pharmacy (IOSRPHR) is an open access online & offline peer reviewed international journal, which publishes innovative research papers, reviews, mini-reviews, short communications and notes dealing with Pharmaceutical Sciences( Pharmaceutical Technology, Pharmaceutics, Biopharmaceutics, Pharmacokinetics, Pharmaceutical/Medicinal Chemistry, Computational Chemistry and Molecular Drug Design, Pharmacognosy & Phytochemistry, Pharmacology, Pharmaceutical Analysis, Pharmacy Practice, Clinical and Hospital Pharmacy, Cell Biology, Genomics and Proteomics, Pharmacogenomics, Bioinformatics and Biotechnology of Pharmaceutical Interest........more details on Aim & Scope).
Flu Vaccine Alert in Bangalore Karnatakaaddon Scans
As flu season approaches, health officials in Bangalore, Karnataka, are urging residents to get their flu vaccinations. The seasonal flu, while common, can lead to severe health complications, particularly for vulnerable populations such as young children, the elderly, and those with underlying health conditions.
Dr. Vidisha Kumari, a leading epidemiologist in Bangalore, emphasizes the importance of getting vaccinated. "The flu vaccine is our best defense against the influenza virus. It not only protects individuals but also helps prevent the spread of the virus in our communities," he says.
This year, the flu season is expected to coincide with a potential increase in other respiratory illnesses. The Karnataka Health Department has launched an awareness campaign highlighting the significance of flu vaccinations. They have set up multiple vaccination centers across Bangalore, making it convenient for residents to receive their shots.
To encourage widespread vaccination, the government is also collaborating with local schools, workplaces, and community centers to facilitate vaccination drives. Special attention is being given to ensuring that the vaccine is accessible to all, including marginalized communities who may have limited access to healthcare.
Residents are reminded that the flu vaccine is safe and effective. Common side effects are mild and may include soreness at the injection site, mild fever, or muscle aches. These side effects are generally short-lived and far less severe than the flu itself.
Healthcare providers are also stressing the importance of continuing COVID-19 precautions. Wearing masks, practicing good hand hygiene, and maintaining social distancing are still crucial, especially in crowded places.
Protect yourself and your loved ones by getting vaccinated. Together, we can help keep Bangalore healthy and safe this flu season. For more information on vaccination centers and schedules, residents can visit the Karnataka Health Department’s official website or follow their social media pages.
Stay informed, stay safe, and get your flu shot today!
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These simplified slides by Dr. Sidra Arshad present an overview of the non-respiratory functions of the respiratory tract.
Learning objectives:
1. Enlist the non-respiratory functions of the respiratory tract
2. Briefly explain how these functions are carried out
3. Discuss the significance of dead space
4. Differentiate between minute ventilation and alveolar ventilation
5. Describe the cough and sneeze reflexes
Study Resources:
1. Chapter 39, Guyton and Hall Textbook of Medical Physiology, 14th edition
2. Chapter 34, Ganong’s Review of Medical Physiology, 26th edition
3. Chapter 17, Human Physiology by Lauralee Sherwood, 9th edition
4. Non-respiratory functions of the lungs https://academic.oup.com/bjaed/article/13/3/98/278874
Lung Cancer: Artificial Intelligence, Synergetics, Complex System Analysis, S...Oleg Kshivets
RESULTS: Overall life span (LS) was 2252.1±1742.5 days and cumulative 5-year survival (5YS) reached 73.2%, 10 years – 64.8%, 20 years – 42.5%. 513 LCP lived more than 5 years (LS=3124.6±1525.6 days), 148 LCP – more than 10 years (LS=5054.4±1504.1 days).199 LCP died because of LC (LS=562.7±374.5 days). 5YS of LCP after bi/lobectomies was significantly superior in comparison with LCP after pneumonectomies (78.1% vs.63.7%, P=0.00001 by log-rank test). AT significantly improved 5YS (66.3% vs. 34.8%) (P=0.00000 by log-rank test) only for LCP with N1-2. Cox modeling displayed that 5YS of LCP significantly depended on: phase transition (PT) early-invasive LC in terms of synergetics, PT N0—N12, cell ratio factors (ratio between cancer cells- CC and blood cells subpopulations), G1-3, histology, glucose, AT, blood cell circuit, prothrombin index, heparin tolerance, recalcification time (P=0.000-0.038). Neural networks, genetic algorithm selection and bootstrap simulation revealed relationships between 5YS and PT early-invasive LC (rank=1), PT N0—N12 (rank=2), thrombocytes/CC (3), erythrocytes/CC (4), eosinophils/CC (5), healthy cells/CC (6), lymphocytes/CC (7), segmented neutrophils/CC (8), stick neutrophils/CC (9), monocytes/CC (10); leucocytes/CC (11). Correct prediction of 5YS was 100% by neural networks computing (area under ROC curve=1.0; error=0.0).
CONCLUSIONS: 5YS of LCP after radical procedures significantly depended on: 1) PT early-invasive cancer; 2) PT N0--N12; 3) cell ratio factors; 4) blood cell circuit; 5) biochemical factors; 6) hemostasis system; 7) AT; 8) LC characteristics; 9) LC cell dynamics; 10) surgery type: lobectomy/pneumonectomy; 11) anthropometric data. Optimal diagnosis and treatment strategies for LC are: 1) screening and early detection of LC; 2) availability of experienced thoracic surgeons because of complexity of radical procedures; 3) aggressive en block surgery and adequate lymph node dissection for completeness; 4) precise prediction; 5) adjuvant chemoimmunoradiotherapy for LCP with unfavorable prognosis.
The prostate is an exocrine gland of the male mammalian reproductive system
It is a walnut-sized gland that forms part of the male reproductive system and is located in front of the rectum and just below the urinary bladder
Function is to store and secrete a clear, slightly alkaline fluid that constitutes 10-30% of the volume of the seminal fluid that along with the spermatozoa, constitutes semen
A healthy human prostate measures (4cm-vertical, by 3cm-horizontal, 2cm ant-post ).
It surrounds the urethra just below the urinary bladder. It has anterior, median, posterior and two lateral lobes
It’s work is regulated by androgens which are responsible for male sex characteristics
Generalised disease of the prostate due to hormonal derangement which leads to non malignant enlargement of the gland (increase in the number of epithelial cells and stromal tissue)to cause compression of the urethra leading to symptoms (LUTS
ARTIFICIAL INTELLIGENCE IN HEALTHCARE.pdfAnujkumaranit
Artificial intelligence (AI) refers to the simulation of human intelligence processes by machines, especially computer systems. It encompasses tasks such as learning, reasoning, problem-solving, perception, and language understanding. AI technologies are revolutionizing various fields, from healthcare to finance, by enabling machines to perform tasks that typically require human intelligence.
MANAGEMENT OF ATRIOVENTRICULAR CONDUCTION BLOCK.pdfJim Jacob Roy
Cardiac conduction defects can occur due to various causes.
Atrioventricular conduction blocks ( AV blocks ) are classified into 3 types.
This document describes the acute management of AV block.
Explore natural remedies for syphilis treatment in Singapore. Discover alternative therapies, herbal remedies, and lifestyle changes that may complement conventional treatments. Learn about holistic approaches to managing syphilis symptoms and supporting overall health.
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Structural Studies of Human GBE1 and Relevance to APBD
1. Structural studies of human
GBE1 and relevance to APBD
Wyatt Yue
Structural Genomics Consortium (SGC)
University of Oxford
APBDRF Meeting Dec 2013
2. • Public-private partnership
• High-throughput structural & chemical
biology of human proteins
• Diverse protein families/biology areas
• Open access research model
Combining structural biology, protein biochemistry
to understand inborn errors of metabolism
www.thesgc.org/wyatt
wyatt.yue@sgc.ox.ac.uk
3. Studying enzyme shapes
• Enzymes are proteins
- building blocks (amino acids)
- have different shapes
• Shapes can change too!
• Understanding enzyme
functions/malfunctions –
need to know its shapes
… seeing molecules in action
4. ‘Taking pictures’ of protein structures
1. make proteins of
interest (expression, purification)
3. take picture!
(x-ray diffraction)
2. arrange them in order
(crystallization)
4. develop the
film (modelling)
6. Glycogen Synthesis
human GYG1 structure
priming
glycogenin
GSD type XV
elongation
Cartoon impression of
glycogen granule
glycogenin
glycogen
synthase
GSD type 0
branching
glycogenin
human GBE1 structure
branching
enzyme
GSD type IV (liver)
Adult Polyglucosan Body Disease
glycogen
synthase
7. Multi-construct approach
1
700 702
E coli structure
63
63
st round
1
70
Insoluble in E. coli 70
79
79
2nd round
Soluble in
insect cells
3rd round
Fine-tune
c000
c001
c002
c003
c004
c005
c101
c102
c103
c104
c105
c106
c107
c108
1
1
16
16
38
38
54
54
L28
P36
Y41
R47
c109
c110
c111
c112
c011, c012
c013, c014
c015, c016
c017, c018
•
•
•
Full length and a series of truncations
N- and C-termini nibbling
N-terminal His6 tagged fusion
8. GBE1 Purification & Crystallization
GBE1A-c014 (recombinant Pro37-Leu700 with a TEV-cleavable His6-tag)
Growth from 4L of Baculovirus-infected insect Sf9 cells
2. Size exclusion
1. Ni affinity
250
FT BB1 WB1
E1
E3
BB2
WB2 E2
150
3. His-tag removal by TEV
4. Second Ni affinity (to
remove uncut protein, TEV)
GBE1A c011 E1 s200 GF 300812:Sample1Title_UV
GBE1A c011 E1 s200 GF 300812:Sample1Title_Logbook
GBE1A c011 E1 s200 GF 300812:Sample1Title_Fractions
u/c
GBE1A c011 E1 s200 GF 300812:Sample1Title_Inject
mAU
350
Pooled
Pooled
200
Pooled
150
Stop Collecting Fractions
15
Start Collecting Fractions
Sample 1 Name
50
Finish Washing Fraction Collector Outlet
100
Wash Fraction Collector Outlet
25
20
BB1 WB
GF
BB2 EB
250
75
37
FT
300
100
50
cut
0
10
A2
0
A4
A6
20
A7
40
A9 A11 B12 B10 B8
B6
60
B4
B2
C1
C3
C5
C7
80
C9 C11 D12 D10 D8
100
D6
D4
D2
E1
E3
E5
120
E7
E9 E11 F12 F10
F8
140
ml
Concentrated, purified
15% PEG 3350
0.15M sodium succinate
9. Structure determination
• Three structure data:
– apo GBE1, GBE1-acarbose and GBE1-Glc7
Complex with
Overall structure sugar chains
Mapping of gbe1 mutations
Y329 site
11. p.Y329S mutant protein purification
Scale up from 6L insect cell culture
Ni affinity
Total Soluble FT WB1 WB2 WB3 WB4 WB5
Test TEV Cleavage
E1
250
150
100
75
50
37
50
37
25
20
15
25
20
15
GBE1A p.Y329S band confirmed by tryptic digestion.
GF, concentrated
uncut cut
E2
250
150
100
75
Concentrated GF Samples
250
150
100
75
50
37
25
20
15
GBE1A-c201 e129 p014 p015 (6L, Baculo)
12. Pharmacological Chaperones
Many metabolic disorders are misfolding defects
Pharmacological Chaperone (PCs)
Rescue of stability and function
Examples of PC development
Disease
Protein
Chaperone
Fabry
GLA
DGJ
Gaucher
GBA
IFG, DNJ
GM1
GLB1
NOEV
GM2
HEXA
Pyrimethamine
Pompe
GAA
DNJ
MPSIII
NAGLU
2AcDNJ,
6AcCAS
Fan et al. 1999; Yu et al. 2007;
Matsuda et al. 2003; Maegawa et al. 2007;
Parenti et al. 2007;Ficko-Blean et al. 2008
Yet, their action is counter-intuitive:
They compete with native substrate/cofactor
Strategy: Develop non-competitive, allosteric PCs?
13. Next Action
• Scale up less soluble proteins
– WT full-length protein
– Mutants Y329S, R515H, R524Q
Different tags/fusion proteins
to improve solubility
• Characterize hits/peptides (Tropak/Kakhlon)
– DSF, limited proteolysis
– Aggregation/unfolding assay
– co-crystallization, ITC
Time, protein consuming
15. Working with rare disease patient groups
Inborn errors of metabolism
Rare cancers & developmental disorders
Membrane proteins & rare diseases
Genomic variation & disease
Alex Bullock
‘Stone man Collaborative projects
syndrome’ Funded personnel
Public engagement
Information
16. What can we do to help APBDRF?
Clinical ‘know-how’
WT/mutant protein
Crystals, 3D Structure
hit finding
xtal soaking
in silico Docking
HT Compound Screening
(Michael Tropak)
in silico Ligand Design
(peptides – Or Kakhlon)
Binding, Biochemistry
DSF, ITC, BLI, co-xtal
dose response, affinity
validation
Cellular Assays
Mode of Action
folding, proteolysis, aggregation
Effects on activity, stability
Test Mutants
rescue stability?
Effects on activity
characterization
patient fibroblast cells
Animal model (e.g. mouse)
enzyme activity
Rescue in vivo?
Goal: To develop compounds into a pharmacological chaperone treatment
17. Example of our In Vitro capabilities
Pathfinder Awards for Orphan Diseases
Cystathionine beta synthase (CBS) deficiency
Aim: look for binders at different pockets/regions as chemical starting point
human CBS structure
Allosteric
domain
(AdoMet)
Recombinant protein
Structural Biology
Catalytic
domain
(PLP, haem)
Fragment DSF
Compound library
Cellular assay
Structurally diverse, drug-like hits
Developed In vitro assays to deconvolute binding modes
In silico docking
Limited proteolysis
kp = 0.381 ± 0.037 min-1
Domain mapping
18. ACKNOWLEDGEMENTS
THE MOB
2013
Thomas McCorvie
Dipali Patel
Jolanta Kopec
Stephanie Oerum
Fiona Fitzpatrick
Sean Froese
Wasim Kiyani
CRYSTALLOGRAPHY
Frank von Delft
Tobias Krojer
2012
BIOTECHNOLOGY
Claire Damerell
Pravin Mahajan
FUNDING PARTNERS
wyatt.yue@sgc.ox.ac.uk
www.thesgc.org/wyatt
The Canadian Institutes for Health Research, the Canada Foundation for Innovation, Genome
Canada, GlaxoSmithKline, Lilly Canada, the Novartis Research Foundation, Pfizer, Takeda, the
Ontario Ministry of Economic Development and Innovation, and the Wellcome Trust.
19. Structure determination
• Three structure data:
– apo GBE1, GBE1-acarbose and GBE1-Glc7
Complex with
Overall structure sugar chains
Mapping of gbe1 mutations
Y329 site
21. Thermal stability as a ligand binding assay
Differential Scanning Fluorimetry (DSF), ‘Tm shift’
WT
Small molecule stabilization? –
right shift towards WT
Niesen et al 2007 Nat Methods
Stabilization by
native ligands –
right shift (+ Tm)
Destabilization due to
mutation – left shift (- Tm)
22. DSF considerations for GBE1
No detectable Tm shift with various sugar analogues
N=2 experiments
- Even at higher concentrations as tested previously
- despite electron density in structures!
- These are not natural ligand but shortened versions
Tm for apo protein varies with experiments
- error margins large, with experiments/preps
- is stability concentration dependent?
Is conformational change required for catalysis?
(This increases likelihood of seeing Tm shift)
- Compounds may not be ‘large’ enough a?
Difference between Full-length vs truncated proteins
(previous) E. coli expressed, FL protein
- Tm varies with constructs
- ->N-terminal may be important for stability/disease?
- may explain lack of detectable binding?
Tm = 41.8 ± 2.1 °C
N=8 experiments
Acarbose (4)
Glc4,Glc7
4-mer peptides
24. Structural analysis of Y329 site
WT
p.Y329S
Y329 forms hydrogen bond between its side-chain and backbone carbonyl of His289.
2.5 Å distance in WT increases to 8.5 Å in p.Y329S.
Virus re-amplified ,
extraction buffer optimised to sodium phosphate based,
Talon (cobalt) used instead of Ni-NTA (Nickel).