Linked Life Data

9753432

Source:http://linkedlifedata.com/resource/pubmed/id/9753432

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
39
pubmed:dateCreated
1998-10-28
pubmed:databankReference
pubmed:abstractText
Pyruvate phosphate dikinase (PPDK) catalyzes the interconversion of ATP, Pi, and pyruvate with AMP, PPi, and PEP in three partial reactions: (1) E + ATP --> E.ATP --> E-PP.AMP, (2) E-PP.AMP + Pi --> E-PP.AMP.Pi --> E-P.AMP.PPi, and (3) E-P + pyruvate --> E-P.pyruvate --> E.PEP. The Clostridium symbiosum PPDK structure consists of N-terminal, central, and C-terminal domains. The N-terminal and central domains catalyze partial reactions 1 and 2 whereas the C-terminal and central domains catalyze partial reaction 3. The goal of the present work is to determine where on the N-terminal domain catalysis of partial reactions 1 and 2 occurs and, in particular, where the Pi binding site is located. Computer modeling studies implicated Arg337 as a key residue for Pi binding. This role was tested by site-directed mutagenesis. The R337A PPDK was shown to be impaired in catalysis of the forward (kcat 300-fold lower) and reverse (kcat 30-fold lower) full reactions. Time courses for the single turnover reactions were measured to show that catalysis of partial reaction 1 is 5-fold slower in the mutant, catalysis of the second partial reaction is 140-fold slower in the mutant, and catalysis of the third partial reaction is unaffected. With the exception of the mutation site, the crystal structure of the R337A PPDK closely resembles the structure of the wild-type protein. Thus, the altered kinetic properties observed for this mutant are attributed solely to the elimination of the interaction between substrate and the guanidinium group of the Arg337 side chain. On the basis of these findings we propose that the Pi binding site is located within the crevice of the PPDK N-terminal domain, at a site that is flanked by the ATP beta-P and the Mg2+ cofactor.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Sep
pubmed:issn
0006-2960
pubmed:author
pubmed:issnType
Print
pubmed:day
29
pubmed:volume
37
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
13463-74
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
1998
pubmed:articleTitle
Location of the phosphate binding site within Clostridium symbiosum pyruvate phosphate dikinase.
pubmed:affiliation
Department of Chemistry and Biochemistry, University of Maryland, College Park 27042, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, U.S. Gov't, Non-P.H.S.