Linked Life Data

11858723

Source:http://linkedlifedata.com/resource/pubmed/id/11858723

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
2002-2-22
pubmed:abstractText
Human fibroblast activation protein (FAP), an integral membrane serine protease, was produced in insect cells as a hexa-His-tagged protein using a recombinant baculovirus expression system. Two isoforms of FAP, glycosylated and nonglycosylated, were identified by Western blotting using an anti-His-tag antibody and separated by lectin chromatography. The glycosylated FAP was purified to near homogeneity using immobilized metal affinity chromatography and was shown to have both postprolyl dipeptidyl peptidase and postgelatinase activities. In contrast, the nonglycosylated isoform demonstrated no detectable gelatinase activity by either zymography or a fluorescence-based gelatinase activity assay. The kinetic parameters of the dipeptidyl peptidase activity for glycosylated FAP were determined using dipeptide Ala-Pro-7-amino-trifluoromethyl-coumarin as the substrate. The k(cat) is 2.0 s(-1) and k(cat)/K(m) is 1.0 x 10(4) M(-1) s(-1) at pH 8.5. The pH dependence of k(cat) reveals two ionization groups with pK(a1) of 7.0 and pK(a2) of 11.0. The pH profile of k(cat)/K(m) yields similar results with pK(a1) 6.2 and pK(a2) 11.0. The neutral pK(a1) is associated with His at the active site. The basic pK(a2) might be contributed from an ionization group that is not involved directly in catalysis, instead associated with the stability of the active site structure.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Mar
pubmed:issn
1046-5928
pubmed:author
pubmed:copyrightInfo
Copyright 2002 Elsevier Science (USA).
pubmed:issnType
Print
pubmed:volume
24
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
274-81
pubmed:dateRevised
2010-11-18
pubmed:meshHeading
pubmed:year
2002
pubmed:articleTitle
Expression, purification, and kinetic characterization of full-length human fibroblast activation protein.
pubmed:affiliation
Applied Biotechnology, The Dupont Pharmaceuticals Company, Experimental Station, Wilmington, Delaware 19880-0336, USA. shaoxian@yahoo.com
pubmed:publicationType
Journal Article