Lecture Notes on Bioinformatics and Proteomics in CISC667 - Prof. Li Liao, Study notes of Computer Science

Download Lecture Notes on Bioinformatics and Proteomics in CISC667 - Prof. Li Liao and more Study notes Computer Science in PDF only on Docsity! CISC636, S08, Lec19, Liao 1 CISC 636 Intro to Bioinformatics (Spring 2008) DNA Microarray, 2d gel, MSMS, yeast 2-hybrid. CISC667, S07, Lec22, Liao 2 Gene expression – How many copies of a gene (its product) is present in the cell? – For experimental reasons, gene expressions are measured by numbers of mRNAs, not directly by proteins. (See Proteomics) – Various cell types are due to different genes expressed. – The difference between diseased (e.g., cancerous) and non-diseased – Diseased cells are often resulted from the abnormal levels of expression of key genes. Faw Image Pseudecolor Ried channel a Connect ta computer Tost Cells Cantral Cells Merged image t Harvest RNA t Proloruhiclier ; ii, Tube ‘henserptns ce aia Labeling { eONA cere gert [rat | eceone® povet oar _gy “Red* Cy3 "“Green™ Pr Cyi "Red" Probes Cy reer obes Pinhole ‘Cormet ta computer = a Excitation Lasers Hytzridize to | Microarray Slite o tas) ——___—_—_—_ Normalization and dala analysis: Microarray slide CISC667, S07, Lec22, Liao ADOTOIMALOVA JO TYNUNOL 9L9¥-SLEr SA5Vd “9) YIBWNN ‘281 SMMOA ‘doce IsNONv AMERICAN SOCIETY FOR MICROBIOLOGY Journal of Bacteriology AUGUST 2000, VOLUME 182, NUMBER 16 Published Twice Monthly CISC667, S07, Lec22, Liao CISC667, S07, Lec22, Liao 7 Applications • Inferring transcription regulatory networks • Understanding correlation between genotype and phenotype • predicting genotype <=> phenotype • Phenotypes: – drug/therapy response – drug-drug interactions for expression – drug mechanism – interacting pathways of metabolism CISC667, S07, Lec22, Liao 10 2D gel electrophoresis – Isoelectric points (first dimension) – Molecular weights (second dimension) Both pI and MW are functions of amino acid sequence of a protein. Some proteins do not resolve well by 2D gels. Issues: • Detection of spots (image processing) • Quantification of each spot • Identification of each spot (Mass Spectrometry) a) d b) c) d) > + e) f) 200kDa ae aae Mw Mw 30kDa 8kDa FIGURE 6.18 * Two-dimensional (2D) gel electrophoresis. Each column (a-c) with a gradation of gray shading represents the isoelectric focusing gel with a pH gradient. a) A mixture of proteins (blue drop) is applied to the isoelectric focusing gel and b) exposed to an electrical current. ¢) Proteins migrate to their isoelectric points (pl) and stop moving. d) This tubular gel is placed on top of a slab polyacrylamide gel that contains SDS and is subjected to electro- phoresis (SDS-PAGE). Proteins migrate into the slab gel according to their molecular weights. Yeast cells were grown in rich media and subjected to 2D gel analysis. Using duplicate isoelec- tric focusing gels, large e) and small f) proteins were analyzed on separate gels. The same spots appear at the bottom of e) and the top of f). Molecular weights are resolved on the Y-axis and pls on the X-axis. Panels e) and f) are from the Swiss 2D database at ExPASy. CISC667, S07, Lec22, Liao 11 a) b) c) d) Separation Activation Mass Analysis MS MS @® HoH lonization Activation Mass Analysis MS lonization Separation Ms Mass Analysis ms lonization Separation Activation CISC667, S07, Lec22, Liao 12 | ap Cell population 1 Cell population 2 Combine 4 Purify target proteins P(30%) From cell population 1 Target protein P(70%) | From cell population 2 Digest, Extract peptides Analyze by MS > & x fl Vi Zi 1 i Lio Mass/charge X: Unphosphorylated peptide e Xp: Phosphorylated peptide X g 2 A BC x D + 3 Xp = 0 we oc FIGURE 6.25 ¢ Quantifying relative levels of phospho- rylation. Bar graphs A, B, C, and D indicate no cifference for proteins A-D in the two populations. Compared to population 1, unphosphorylated protein X was reduced in population 2. Phosphorylated protein X (Xp) was increased in population 2 CISC667, S07, Lec22, Liao 15 Unclear classification lonic homeostasis Cellular organization Unclassified proteins Metabolism Cell rescue, cell defense / \ Energy and cell aging ; Signal transduction _ Cell growth, division, Cellular biogenesis a DNA synthesis Intracellular transport Transport facilitation Protein destination Transcription Protein synthesis FIGURE 6.9 ® Bar code analysis of biological processes. Distribution of functional classes of essential (inner circle) and nonessential (outer circle) genes using criteria from the Munich Information Center for Protein Sequences (MIPS). CISC667, S07, Lec22, Liao 16  Dep1 Lsm5 cf sm lems YLR269C FIGURE 6.15 ¢ Proteomics circuit showing the interactions of RNA splicing pro- teins. The proteins are indicated by large blue nodes and their interactions with lines. The dots on the lines help you follow each line. Blue lines show interactions that were detected by traditional Y2H array screens, black from multiple high-throughput screens, and gray from literature and array screens. The black arrows point away from the protein used as bait in the screens. Small gray nodes indicate other protein-protein interactions not highlighted here. CISC667, S07, Lec22, Liao 17 * Gald protein: the two domains of the proteim do not need to be transcribed in a single protein * Just as long as they come to interact Two other protein domains inter CISC667, S07, Lec22, Liao 20  asm CISC667, S07, Lec22, Liao 21 Screened against Screened against A AN Screened against Screened against Screened against 1 CISC667, S07, Lec22, Liao Screened against 22