Simulation studies connecting protein structure to cellular fitness

Abstract

Mutations in protein sequences affect the 3D structures of proteins, with consequences for the fitness of the organism. The fitness effects of mutations in turn determine how a protein may evolve. We report studies that combine biophysical modeling of fitness, molecular simulation, and bioinformatic or experimental data to find relationships between protein structure and cellular fitness. In Chapter 2, we examine how proteins that differ in native structure differ in their likelihood to evolve cotranslational folding. Cotranslational protein folding is the formation of native structure during the biological process of protein translation. We use simple model proteins to examine how proteins with different degrees of local and non-local native contacts evolve under a scenario where folding kinetics and stability of a protein determines organismic fitness. We find that proteins with more local contacts in their native structures evolve a cotranslational mechanism of folding, whereas proteins with non-local native contacts delay folding during translation to fold all at once toward the end of translation. In Chapter 3, we move from model proteins to studying replacement of an α-helix in E. coli dihydrofolate reductase (DHFR). We examine what factors determine local structure in a segment of a protein. Evolutionary analysis and molecular dynamics simulation are used to study the loss in catalytic activity and fitness exhibited by the DHFR chimeras. Structurally, we find that high activity chimeras maintain a wildtype-like helical conformation, and we explain our results in terms of biophysical compatibility of the replacement segment. Finally, in Chapter 4, we describe the use of atomistic Monte Carlo protein simulation to characterize alternative disulfide linkages in E. coli chaperone protein HdeA. HdeA is an 89-residue, acid-activated periplasmic chaperone protein that forms a folded homodimer in the inactive state. HdeA natively has a disulfide bond between cysteine residues 18 and 66. A deep mutational scanning assay was performed to characterize the fitness effects of altering the positioning of the two cysteine residues in HdeA. Expression of certain, deleterious double cysteine variants causes cell lysis. In simulations of HdeA variants with non-native disulfide linkages, we observe an alternative dimeric conformation for HdeA. We further find associations between simulation observables and experimentally measured fitness that suggest deleterious cysteine variants have more disordered conformations.