The ability of proteins to change structure impacts upon many biological processes, including the evolution of new folds and functions from ancestral precursors, the regulation of allosteric systems, and the development of amyloid diseases. The long-term goal of this work is to understand how large-scale structural change is promoted or prevented by changes in amino-acid sequence. For this purpose, I have chosen as a model system the cro family of bacteriophage DNA-binding proteins. Members of this family have related sequences but very different folds. Lambda cro has a dimeric alpha+beta fold, while preliminary results show that P22 cro has a monomeric all-alpha fold consisting of five helices. The lambda cro variant A33W/F58D appears to represent still a third, intermediate type of structure, a monomeric alpha+beta fold. The sequences of these proteins have been aligned, and are homologous even in the region where their structures are most different. I intend to use lambda cro, P22 cro and lambda cro A33W/F58D as a representative set of aligned sequences encoding three distinct structures. Using these approaches, I will a) elucidate the key sequence determinants which distinguish the all-alpha and alpha+beta folds in cro proteins, b) elucidate the sequence determinants of strong dimerization in lambda cro, and characterize the structure of intermediate monomeric alpha+beta variants, c) design structurally ambivalent sequences which are compatible with both five-helix and alpha+beta folds. In addition, I will characterize additional proteins in this family to expand the system beyond the P22/lambda model.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
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Molecular and Cellular Biophysics Study Section (BBCA)
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Wehrle, Janna P
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University of Arizona
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Stewart, Katie L; Rathore, Deepali; Dodds, Eric D et al. (2018) Increased sequence hydrophobicity reduces conformational specificity: A mutational case study of the Arc repressor protein. Proteins :
Kumirov, Vlad K; Dykstra, Emily M; Hall, Branwen M et al. (2018) Multistep mutational transformation of a protein fold through structural intermediates. Protein Sci 27:1767-1779
Eaton, Karen V; Anderson, William J; Dubrava, Matthew S et al. (2015) Studying protein fold evolution with hybrids of differently folded homologs. Protein Eng Des Sel 28:241-50
Stewart, Katie L; Dodds, Eric D; Wysocki, Vicki H et al. (2013) A polymetamorphic protein. Protein Sci 22:641-9
Stewart, Katie L; Nelson, Michael R; Eaton, Karen V et al. (2013) A role for indels in the evolution of Cro protein folds. Proteins 81:1988-96
Dodds, Eric D; Blackwell, Anne E; Jones, Christopher M et al. (2011) Determinants of gas-phase disassembly behavior in homodimeric protein complexes with related yet divergent structures. Anal Chem 83:3881-9
Bouvignies, Guillaume; Vallurupalli, Pramodh; Cordes, Matthew H J et al. (2011) Measuring 1HN temperature coefficients in invisible protein states by relaxation dispersion NMR spectroscopy. J Biomol NMR 50:13-8
Anderson, William J; Van Dorn, Laura O; Ingram, Wendy M et al. (2011) Evolutionary bridges to new protein folds: design of C-terminal Cro protein chameleon sequences. Protein Eng Des Sel 24:765-71
Bouvignies, Guillaume; Korzhnev, Dmitry M; Neudecker, Philipp et al. (2010) A simple method for measuring signs of (1)H (N) chemical shift differences between ground and excited protein states. J Biomol NMR 47:135-41
Dubrava, Matthew S; Ingram, Wendy M; Roberts, Sue A et al. (2008) N15 Cro and lambda Cro: orthologous DNA-binding domains with completely different but equally effective homodimer interfaces. Protein Sci 17:803-12

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