Protein Science
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Protein Science (2007), 16:1609-1616. Published by Cold Spring Harbor Laboratory Press. Copyright © 2007 The Protein Society
This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Johnson, R. J.
Right arrow Articles by Raines, R. T.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Johnson, R. J.
Right arrow Articles by Raines, R. T.
Social Bookmarking
Add to CiteULike   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati  
What's this?

Genetic selection reveals the role of a buried, conserved polar residue

R. Jeremy Johnson1,3, Shawn R. Lin1,3, and Ronald T. Raines1,2

1 Department of Biochemistry, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA
2 Department of Chemistry, University of Wisconsin-Madison, Madison, Wisconsin 53706, USA

(RECEIVED April 10, 2007; FINAL REVISION May 14, 2007; ACCEPTED May 15, 2007)

The burial of nonpolar surface area is known to enhance markedly the conformational stability of proteins. The contribution from the burial of polar surface area is less clear. Here, we report on the tolerance to substitution of Ser75 of bovine pancreatic ribonuclease (RNase A), a residue that has the unusual attributes of being buried, conserved, and polar. To identify variants that retain biological function, we used a genetic selection based on the intrinsic cytotoxicity of ribonucleolytic activity. Cell growth at 30°C, 37°C, and 44°C correlated with residue size, indicating that the primary attribute of Ser75 is its small size. The side-chain hydroxyl group of Ser75 forms a hydrogen bond with a main-chain nitrogen. The conformational stability of the S75A variant, which lacks this hydrogen bond, was diminished by {Delta}{Delta}G = 2.5 kcal/mol. Threonine, which can reinstate this hydrogen bond, provided a catalytically active RNase A variant at higher temperatures than did some smaller residues (including aspartate), indicating that a secondary attribute of Ser75 is the ability of its uncharged side chain to accept a hydrogen bond. These results provide insight on the imperatives for the conservation of a buried polar residue.

Keywords: conformational stability; genetic selection; hydrogen bond; hydrophobic core; molecular evolution; ribonucleases


Add to CiteULike CiteULike   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati    What's this?




HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Copyright © 2007 by The Protein Society.