Linked Life Data

12646374

Source:http://linkedlifedata.com/resource/pubmed/id/12646374

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
1-3
pubmed:dateCreated
2003-3-20
pubmed:abstractText
The catalytically inactive mutant S195A was used to study the interaction of thrombin with substrates under equilibrium conditions. By monitoring changes in intrinsic fluorescence, we measured dissociation constants for a variety of synthetic substrates, PAR peptides and the inhibitor PPACK. The S195A mutant retains the Na(+)-binding properties of the wild type, and substrate binding to the mutant is enhanced by the presence of Na(+). Temperature dependence studies allowed calculation of the thermodynamic parameters of substrate binding at the active site and showed a negligible deltaC(p). Titration of synthetic substrates carrying substitutions at the P1-P3 positions revealed energetics consistent with the specificity hierarchy identified in hydrolysis by the wild type. Titration with PAR peptides, which interact with both the active site and exosite I of thrombin, also showed consistency with the results obtained with the wild type at steady state. These findings demonstrate that inactive mutants of enzymes make it possible to dissect the equilibrium components linked to substrate binding and complement information on the kinetic properties of the wild type.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:issn
0301-4622
pubmed:author
pubmed:issnType
Print
pubmed:volume
100
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
315-23
pubmed:dateRevised
2008-11-21
pubmed:meshHeading
pubmed:year
2003
pubmed:articleTitle
Dissecting substrate recognition by thrombin using the inactive mutant S195A.
pubmed:affiliation
Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, Box 8231, St. Louis, MO 63110, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S.