The Complex Role of Sequence and Structure in the Stability and Function of the TIM Barrel Proteins
Authors
Chan, Yvonne H.Faculty Advisor
C. Robert Matthews; Konstantin B. ZeldovichAcademic Program
Biochemistry and Molecular PharmacologyUMass Chan Affiliations
Biochemistry and Molecular PharmacologyDocument Type
Doctoral DissertationPublication Date
2017-11-03Keywords
TIM barrelIGPS
protein stability
evolution
mutation
biophysics
fitness landscapes
protein folding
Biochemistry
Biophysics
Molecular Biology
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Show full item recordAbstract
Sequence divergence of orthologous proteins enables adaptation to a plethora of environmental stresses and promotes evolution of novel functions. As one of the most common motifs in biology capable of diverse enzymatic functions, the TIM barrel represents an ideal model system for mapping the phenotypic manifestations of protein sequence. Limits on evolution imposed by constraints on sequence and structure were investigated using a model TIM barrel protein, indole-3-glycerol phosphate synthase (IGPS). Exploration of fitness landscapes of phylogenetically distant orthologs provides a strategy for elucidating the complex interrelationship in the context of a protein fold. Fitness effects of point mutations in three phylogenetically divergent IGPS proteins during adaptation to temperature stress were probed by auxotrophic complementation of yeast with prokaryotic, thermophilic IGPS. Significant correlations between the fitness landscapes of distant orthologues implicate both sequence and structure as primary forces in defining the TIM barrel fitness landscape. These results suggest that fitness landscapes of point mutants can be successfully translocated in sequence space, where knowledge of one landscape may be predictive for the landscape of another ortholog. Analysis of a surprising class of beneficial mutations in all three IGPS orthologs pointed to a long-range allosteric pathway towards the active site of the protein. Biophysical and biochemical analyses provided insights into the molecular mechanism of these beneficial fitness effects. Epistatic interactions suggest that the helical shell may be involved in the observed allostery. Taken together, knowledge of the fundamental properties of the TIM protein architecture will provide new strategies for de novo protein design of a highly targeted protein fold.DOI
10.13028/M2J09HPermanent Link to this Item
http://hdl.handle.net/20.500.14038/32317Rights
Licensed under a Creative Commons licenseDistribution License
http://creativecommons.org/licenses/by/4.0/ae974a485f413a2113503eed53cd6c53
10.13028/M2J09H
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