Linked Life Data

7827038

Source:http://linkedlifedata.com/resource/pubmed/id/7827038

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
3
pubmed:dateCreated
1995-2-23
pubmed:abstractText
Aspartic acid 97 in the inorganic pyrophosphatase of Escherichia coli (E-PPase) has been identified as an evolutionarily conserved residue forming part of the active site [Cooperman et al. (1992) Trends Biochem. Sci. 17, 262-266]. Here we determine the effect of D97E substitution on several kinetic and thermodynamic properties of E-PPase, including rate and equilibrium constants for enzyme-catalyzed PPi.Pi equilibration at pH 7.2 and 8.0, Mg2+ affinity in the presence and absence of substrate, and the Mg2+ and pH dependence of kcat and Km. We find the major effects of D97E substitution are to (a) decrease markedly the pH-independence rates of both PPi hydrolysis and, especially, PPi resynthesis on the enzyme, (b) selectively destabilize both the EMg4PPi complex and the transition state between this complex and the EMg2(MgPi)2 complex, (c) raise the pKa of the basic group "essential" for PPi hydrolysis and for productive PPi binding by 1.5 and > 2.2 log units, respectively, (d) distort a site to which Mg2+ binds in the absence of substrate such that occupancy of the site by Mg2+ no longer confers enzymatic activity, and (e) decrease the affinity of one of the two Mg2+ ions that binds to enzyme in the presence of substrate. That this multiplicity of effects arises from a single Asp to Glu substitution suggests, in the absence of any evidence for a generalized structural change, a tightly integrated active site in which the perturbation induced by conservative substitution at a single location can have widespread functional effects.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jan
pubmed:issn
0006-2960
pubmed:author
pubmed:issnType
Print
pubmed:day
24
pubmed:volume
34
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
792-800
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
1995
pubmed:articleTitle
Effect of D97E substitution on the kinetic and thermodynamic properties of Escherichia coli inorganic pyrophosphatase.
pubmed:affiliation
Department of Chemistry, University of Pennsylvania, Philadelphia 19104-6323.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S.