Lecture Notes - 2 7.24/7.88J/5.48J The Protein Folding Problem Handouts: An Anfinsen paper Reading List Anfinsen Experiments The Problem of the title refers to how the amino acid sequence of a polypeptide chain determines the folded three-dimensional organization of the chain: • Does the sequence determine the structure?? • Protein Denaturation • Emergence of the Problem • Ribonuclease A • Refolding of ribonuclease in vitro. A. Protein Denaturation/InactivationOne of the features that identified proteins as distinctive polymers was 1) Unusual phase transition in semi purified proteinsWhen exposed to relatively gentle conditions outside the range of physiological • heat • pH • salt • organic solvents, e.g. alcohols Lets consider the familiar transition I mentioned last week; • Heat denaturation; cooking the white of an egg • Acid denaturation; Milk turning sour Macroscopic changes in bulk solution; scatters visible light, increase in viscosity - white of egg: >>heat; opaque, hard; cool down; no change: Coagulation, Aggregation, precipitation, denaturation: One of the components is egg white lysozyme: activity assay; hydrolysis of bacterial cell walls: Activity remaining vs. temperature on same axesThis transition: is Denaturation. These transitions were in general found to be irreversible: activity was not recovered upon cooling: Historically the study of the folding and unfolding of proteins emerged from trying to understand this unusual change of state or phase transitions.