Linked Life Data

1655733

Source:http://linkedlifedata.com/resource/pubmed/id/1655733

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
27
pubmed:dateCreated
1991-10-29
pubmed:abstractText
We have identified a binding site for tissue inhibitors of metalloproteinases 2 (TIMP-2) on human 72-kDa gelatinase that is distinct from the active site. 72-kDa progelatinase is found in a complex with TIMP-2 in the medium of cultured cells and can be activated with organomercurial compounds to yield a gelatinolytic proteinase that remains bound to TIMP-2. Removal of TIMP-2 from 72-kDa progelatinase by reverse-phase high performance liquid chromatography, followed by reconstitution of the progelatinase in neutral pH buffer, results in autocatalytic activation. When samples of autoactivated gelatinase were blotted onto nitrocellulose, then probed with 125I-TIMP-2, we found a 29-kDa peptide that was capable of binding TIMP-2. We isolated this fragment and identified it as the region of gelatinase from amino acid 414 to the carboxyl terminus in the primary amino acid sequence of progelatinase. This portion of the molecule does not contain the putative zinc- or gelatin-binding sites and is proteolytically inactive. Incubation of 125I-TIMP-2 with 72-kDa progelatinase-TIMP-2 complexes resulted in a concentration-dependent exchange of labeled TIMP-2 with unlabeled TIMP-2, in both the presence and absence of the metalloproteinase inhibitor 1,10-phenanthroline. Saturation binding kinetics for the active site of 72-kDa gelatinase were measured in pools of the 43-kDa active fragment that results from the autoactivation of 72-kDa progelatinase; this fragment has no carboxyl-terminal TIMP-2 binding capability. Binding of 125I-TIMP-2 to the active site was completely inhibited by 1,10-phenanthroline. Binding kinetics for the putative stabilization site were determined with isolated 72-kDa progelatinase. In the presence of 1,10-phenanthroline, 72-kDa progelatinase bound 125I-TIMP-2 but not 125I-TIMP-1. Scatchard analysis yielded an approximate dissociation constant (Kd) of 0.72 nM for the active site and 0.42 nM for the stabilization site.
pubmed:grant
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Sep
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
25
pubmed:volume
266
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
17972-7
pubmed:dateRevised
2007-11-14
pubmed:meshHeading
pubmed:year
1991
pubmed:articleTitle
Binding of tissue inhibitor of metalloproteinases 2 to two distinct sites on human 72-kDa gelatinase. Identification of a stabilization site.
pubmed:affiliation
Laboratory of Radiobiology and Environmental Health, University of California, San Francisco 94143-0750.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S., Research Support, U.S. Gov't, Non-P.H.S.