Table of Contents
Bio 101: Genomics & Computational Biology
Protein1: Last week's take home lessons
Protein2: Today's story & goals
Why purify?
(Protein) Purification Methods
Protein Separation by Gel Electrophoresis
Comparison of predicted with observed protein properties (localization, postsynthetic modifications) E.coli
Computationally checking proteomic data
Protein2: Today's story & goals
Cell fraction: Periplasm 2D gel: SDS mobility isoelectic pH
Cell localization predictions
Isotope calculations
Computationally checking proteomic data
High Performance Liquid Chromatography
Mobile Phase of HPLC
Stationary Phase of HPLC
RP-LC calculated observed
PPT Slide
PPT Slide
Directed Analysis of Large Protein Complexes by 2D separation: strong cation exchange and reversed-phased liquid chromatography.
A new 40S subunit protein
Protein2: Today's story & goals
PPT Slide
Tandem Mass Spectrometry
Ions
Peptide Fragmentation and Ionization
Tandem Mass Spectra Analysis
Mass Spectrum Interpretation Challenge
A dynamic programming approach to de novo peptide sequencing via tandem mass spectrometry
SEQUEST: Sequence-Spectrum Correlation
Protein2: Today's story & goals
Expression quantitation methods
Molecules per cell
MS Protein quantitation R=.84
MS quantitation reproducibility
Correlation between protein and mRNA abundance in yeast
Normality tests
Spearman correlation rank test
Correlation of (phosphorimager 35S met) protein & mRNA
Observed (Phosphorimage) protein levels vs. Codon Adaptation Index (CAI)
ICAT Strategy for Quantifying Differential Protein Expression.
Mass Spectrum and Reconstructed Ion Chromatograms.
Protein & mRNA Ratios +/- Galactose
Protein2: Today's story & goals
Post-synthetic modifications
32P labeled phoshoproteomics
Natural crosslinks
Crosslinked peptide Matrix-assisted laser desorption ionization Post-Source Decay (MALDI-PSD-MS)
Constraints for homology modeling based on MS crosslinking distances
Homology modeling accuracy
Top 20 threading models for FGF ranked by crosslinking constraint error
Protein2: Today's story & goals
Challenges for accurately measuring metabolites
Databases
Y= RP LC retention time in min. (higher hydro- phocity) X = Mass
Metabolite fragmentation & stable isotope labeling
Isotopomers
MetaFoR: Metabolic Flux Ratios
A functional genomics strategy that uses metabolome data to reveal the phenotype of silent mutations
Types of interaction models
How do enzymes & substrates formally differ?
Enzyme rate equations with one Substrate & one Product
Enzyme Kinetic Expressions
Human Red Blood Cell ODE model
Red Blood Cell in Mathematica
Protein2: Today's story & goals
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