Proteomics

1. Proteomics • The term indicates the proteins expressed by a genome. • Expression proteomics, cell map proteomics, structural proteomics. • Proteome analysis means study of modification of proteins in different stages of cell growth---there are five main steps: KK,DBT, ANU

2. Sample collection and storage, handling • Separation by 2D PAGE • Identification by mass • Characterization • Storage, manipulation and comparison of data using BI tools. • Tools of analysis:Microarray • Mass spectrometry approach

3. Protein interaction study • Non homology methods • Domain fusion • Conservation of gene position • Phylogenetic profiles • All-against-all comparison • Cluster analysis • Clustering by single linkage • Core proteome

4. Metabolic pathways • Metabolic networks-KEGG • 2 sections—met path section and regu path section • Genetic networks and their analysis facilitate pathway analysis. • Ligand, drug, pathway, enzyme studies are done. • WIT, BRITE, CSNDB, SPAD.

5. Protein structure Prediction Levels of protein structure— Primary Secondary Tertiary (protein folding) Quartenary

6. Protein structure prediction has certain constrains like mean force potential derived from database, generates incorrect models---inter-residue contacts, so on • There are several problems related to protein folding. • The prediction is done as ID, 2D, 3D.

7. Determinants of protein structure • Hierarchy of protein • Sequence determinants • Secondary structure • Function specific motifs • Classification of proteins based on structure • Components of stability, mutations,,,,,,

8. Known and Unknown fold structure prediction • The first step of structure prediction starts with sequence alignment. • The process to generate a model needs a unknown structure (U) and template with known structure (T). • The sequences having 70-90% similarity generate accurate models, the process is homology modelling

9. The sequences which share less than 30% similarity are modeled by remote homology modeling called threading. • Steps of modeling for >30% –Database search, alignment , multiple alignment, modeling tools MODELLER-Swiss-Model • Steps of threading—remote homologue screening, alignment, modeling • Profile method and core threading model are algorithms used in threading. • Threading tools—PHDthreader, T3P2

10. Methods of structure prediction for unknown folds • Prediction in 1D—Solvent accessibility prediction, predicting transmembrane helices,identification and characterization-composition using ExPASy, some of the other identification tools like pepMAPPER, Mascot, Pepsea, PROWL---Primary structure analysis—tools for physical properties

11. Secondary strc prediction • Primary structure analysis by SAP-statistical analysis of protein sequences. • Tools for physical properties study-ex: Protpram, PEST identifier, multicoil, paircoil,,,, • Study of profiles, motifs and fingerprints. • Algorithms used in sec structure prediction—chou-fasman and GOR methods, NJ method and HMM. • Tools-nnpredict, PREDATOR, GORI, PSA,,,

12. Tertiary strc prediction • Called prediction in 3D • The method used is ab initio prediction where template is not required and based on mean force potentials the models are generated. • Tools-Swiss model, Geno3D, SWEET,,, • DB-3D ALI, CATH, PDB, MMDB, PIR, SCOP,,,,

13. Protein function prediction • Protein seq dtns struc and that dtns function. • At three levels the prediction is done ie., first , second and third generation using different tools. • Accuracy of prediction is given by a index: Qipred=Pi/(Pi+Oi) Pi no of residues in the I state(actual prediction).

14. Oi is the number of residues predicted but not actual. • Qi abs= Pi/ (Pi+Ui) • Ui is the no of residues actually in I state but not predicted, Qi abs is the index of accuracy • Q3= (Palpha+Pbeta+Pcoil)/T • Q3 is global accuracy it is 33% • Applying Matthews coefficients we derive an equation

15. Ci= (PiNi-UiOi)/ [(Pi+Ui)(Pi+Oi)(Ni+Ui)(Ni+Oi)]rr1/2 • Based on this equation the system does a random calcualtion.