Font Size: a A A

X-ray crystallographic studies of lysostaphin ALE-1, HCV NS3 helicase, and staphylococcal nuclease

Posted on:2003-01-28Degree:Ph.DType:Dissertation
University:University of ArkansasCandidate:Lu, ZhiqiangFull Text:PDF
GTID:1460390011485374Subject:Chemistry
Abstract/Summary:
Lysostaphin is a peptidoglycan hydrolase that specifically lyses S. aureus cell walls by cleaving the pentaglycine linkage between the peptidoglycan chains. Its C-terminal targeting domain, a member of the newly identified bacterial SH3b family, directs the enzyme to a specific site on the cell wall. The crystal structure of the targeting domain was solved to 1.75A resolution. It is structurally similar to typical eukaryotic SH3 domains despite the low sequence similarity; however, significant structural changes at the loop regions are responsible for eliminating the capability of SH3b for binding proline-rich substrates. The targeting domain also displays high sequence homology to other SH3b proteins that target gram-positive cell walls. Besides their structural roles, these conserved residues form a large hydrophobic surface area that can potentially interact with the common features of the gram-positive cell wall. The N-terminus of the targeting domain is highly conserved among proteins targeting S. aureus cell walls, suggesting that the N-terminal region is required for achieving substrate specificity.;In vitro biological studies have shown that Hepatitis C Virus NS3 helicase is active as an oligomer; however, the previously reported oligonucleotide (dU)8-bound helicase structure (1A1V.pdb) supports a monomeric "open-closed" mechanism for its translocation along the nucleic acid. The crystal structure of HCV helicase was determined in the presence of an oligonucleotide (dU)16. A dimeric helicase-oligonucleotide complex was found in the asymmetric unit, displaying two slightly different nucleic acid binding modes. The dimeric helicase structure does not challenge the "open-closed" model or the more general "inch-worm" mechanism; instead, it may represent a modified "inch-worm" mechanism in which helicases function cooperatively in the oligomeric form.;Crystal structures of wild-type and mutant staphylococcal nucleases (SNases) were solved to various resolutions and mutant structures were compared with that of wild-type. Mutations on the wild-type SNase alter the protein stability and the structure simultaneously. Changes in van der Waals contacts, Voronoi volume of the hydrophobic core, and hydrogen bonds, were carefully examined; the repacking of the hydrophobic core was also observed in some of the mutant structures. To some extent, specific structural changes could be correlated to the changes in stability.
Keywords/Search Tags:Helicase, Cell walls, Structure, Crystal, Targeting domain, Changes
Related items