Linked Life Data

8999955

Source:http://linkedlifedata.com/resource/pubmed/id/8999955

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
4
pubmed:dateCreated
1997-2-21
pubmed:abstractText
The bifunctional glutathionylspermidine synthetase/amidase from Escherichia coli catalyzes both the ATP-dependent formation of an amide bond between N1 of spermidine (N-(3-amino)propyl-1, 4-diaminobutane) and the glycine carboxylate of glutathione (gamma-Glu-Cys-Gly) and the opposing hydrolysis of this amide bond (Bollinger, J. M., Jr., Kwon, D. S., Huisman, G. W., Kolter, R., and Walsh, C. T. (1995) J. Biol. Chem. 270, 14031-14041). In our previous work describing its initial characterization, we proposed that the 619-amino acid (70 kDa) protein might possess separate amidase (N-terminal) and synthetase (C-terminal) domains. In the present study, we have confirmed this hypothesis by expression of independently folding and functional amidase and synthetase modules. A fragment containing the C-terminal 431 amino acids (50 kDa) has synthetase activity only, with steady-state kinetic parameters similar to the full-length protein. A fragment containing the N-terminal 225 amino acids (25 kDa) has amidase activity only and is significantly activated relative to the full-length protein for hydrolysis of glutathionylspermidine analogs. This observation suggests that the amidase activity in the full-length protein is negatively autoregulated. The amidase active site catalyzes hydrolysis of amide and ester derivatives of glutathione (e.g. glutathione ethyl ester and glutathione amide) but lacks activity toward acetylspermidine (N1 and N8) and acetylspermine (N1), indicating that glutathione provides the primary recognition determinants for glutathionylspermidine amide bond cleavage. No metal ion is required for the amidase activity. A tetrahedral phosphonate analogue of glutathionylspermidine, designed as a mimic of the proposed tetrahedral intermediate for either reaction, inhibits the synthetase activity (Ki approximately 10 microM) but does not inhibit the amidase activity.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jan
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
24
pubmed:volume
272
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
2429-36
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
1997
pubmed:articleTitle
Dissection of glutathionylspermidine synthetase/amidase from Escherichia coli into autonomously folding and functional synthetase and amidase domains.
pubmed:affiliation
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, Non-U.S. Gov't