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Type of Document	Dissertation
Author	Sarisky, Catherine Ann
URN	etd-05272005-121337
Persistent URL	http://resolver.caltech.edu/CaltechETD:etd-05272005-121337
Title	Exploration of the determinants of protein structure and stability by protein design
Degree	PhD
Option	Chemistry
Advisory Committee	Advisor Name Title Richard W. Roberts Committee Chair Douglas C. Rees Committee Member Peter B. Dervan Committee Member Stephen L. Mayo Committee Member
Keywords	gb1 protein design protein structure dead end elimination
Date of Defense	2005-04-27
Availability	unrestricted
Abstract Optimization of Rotamers by Iterative Techniques (ORBIT) has been used to calculate novel sequences for several small proteins. A partial sequence design (20 of 28 residues) is described for the zinc finger Zif268 (beta-beta-alpha) motif. The designed peptide folds without a metal cofactor, despite its small size and the avoidance of the disulfides and unnatural amino acids that are often used to stabilize peptide structures. The utility of ORBIT for predicting the relative stabilities of a series of beta-beta-alpha peptides was investigated. A good correlation between theoretical and experimental stabilities was observed except when the turn residues were changed. This observation led to the discovery that some of these peptides had an unexpected turn conformation. This information was used to design a peptide that is more stable than the original peptide sequence produced with ORBIT. The tolerance of ORBIT for altered backbone coordinates was investigated using the protein domain G-beta1. It was determined that altering the coordinates of the backbone template used in ORBIT altered the sequences selected, but that the fold did not change as a result. The G-beta1 domain was also used to parameterize a methionine inclusion penalty, allowing the inclusion of methionine in ORBIT design calculations while preventing indiscriminate inclusion of methionine at sites where a less flexible side-chain will fit. Lastly, some preliminary work on using ORBIT to design DNA binding interfaces is discussed.
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