PMID- 10024513
OWN - NLM
STAT- MEDLINE
DCOM- 19990506
LR  - 20181113
IS  - 0264-6021 (Print)
IS  - 1470-8728 (Electronic)
IS  - 0264-6021 (Linking)
VI  - 338 ( Pt 2)
IP  - Pt 2
DP  - 1999 Mar 1
TI  - Comparison of the fibrin-binding activities in the N- and C-termini of 
      fibronectin.
PG  - 375-86
AB  - Fibronectin (Fn) binds to fibrin in clots by covalent and non-covalent 
      interactions. The N- and C-termini of Fn each contain one non-covalent 
      fibrin-binding site, which are composed of type 1 (F1) structural repeats. We 
      have previously localized the N-terminal site to the fourth and fifth F1 repeats 
      (4F1.5F1). In the current studies, using proteolytic and recombinant proteins 
      representing both the N- and C-terminal fibrin-binding regions, we localized and 
      characterized the C-terminal fibrin-binding site, compared the relative 
      fibrin-binding activities of both sites and determined the contribution of each 
      site to the fibrin-binding activity of intact Fn. By fibrin-affinity 
      chromatography, a protein composed of the 10F1 repeat through to the C-terminus 
      of Fn (10F1-COOH), expressed in COS-1 cells, and 10F1-12F1, produced in 
      Saccharomyces cerevisiae, displayed fibrin-binding activity. However, since 10F1 
      and 10F1.11F1 were not active, the presence of 12F1 is required for fibrin 
      binding. A proteolytic fragment of 14.4 kDa, beginning 14 residues N-terminal to 
      10F1, was isolated from the fibrin-affinity matrix. Radio-iodinated 14.4 kDa 
      fibrin-binding peptide/protein (FBP) demonstrated a dose-dependent and saturable 
      binding to fibrin-coated wells that was both competitively inhibited and reversed 
      by unlabelled 14.4 kDa FBP. Comparison of the fibrin-binding affinities of 
      proteolytic FBPs from the N-terminus (25.9 kDa FBP), the C-terminus (14.4 kDa) 
      and intact Fn by ELISA yielded estimated Kd values of 216, 18 and 2.1 nM, 
      respectively. The higher fibrin-binding affinity of the N-terminus was 
      substantiated by the ability of both a recombinant 4F1.5F1 and a monoclonal 
      antibody (mAb) to this site to maximally inhibit biotinylated Fn binding to 
      fibrin by 80%, and by blocking the 90% inhibitory activity of a polyclonal 
      anti-Fn, by absorption with the 25.9 kDa FBP. We propose that whereas the 
      N-terminal site appears to contribute to most of the binding activity of native 
      Fn to fibrin, the specific binding of the C-terminal site may strengthen this 
      interaction.
FAU - Rostagno, A A
AU  - Rostagno AA
AD  - Department of Pathology, New York University Medical Center, 400 East 34th 
      Street, New York, NY 10016, USA.
FAU - Schwarzbauer, J E
AU  - Schwarzbauer JE
FAU - Gold, L I
AU  - Gold LI
LA  - eng
GR  - AI 32110/AI/NIAID NIH HHS/United States
GR  - CA 44627/CA/NCI NIH HHS/United States
PT  - Comparative Study
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - England
TA  - Biochem J
JT  - The Biochemical journal
JID - 2984726R
RN  - 0 (DNA Primers)
RN  - 0 (Fibronectins)
RN  - 0 (Peptide Fragments)
RN  - 0 (Recombinant Proteins)
RN  - 9001-31-4 (Fibrin)
SB  - IM
MH  - Amino Acid Sequence
MH  - Animals
MH  - Base Sequence
MH  - COS Cells
MH  - Chromatography, Affinity
MH  - DNA Primers
MH  - Fibrin/*metabolism
MH  - Fibronectins/chemistry/*metabolism
MH  - Humans
MH  - Molecular Sequence Data
MH  - Peptide Fragments/chemistry/isolation & purification/metabolism
MH  - Protein Binding
MH  - Recombinant Proteins/chemistry/isolation & purification/metabolism
PMC - PMC1220063
EDAT- 1999/02/20 00:00
MHDA- 1999/02/20 00:01
PMCR- 1999/09/01
CRDT- 1999/02/20 00:00
PHST- 1999/02/20 00:00 [pubmed]
PHST- 1999/02/20 00:01 [medline]
PHST- 1999/02/20 00:00 [entrez]
PHST- 1999/09/01 00:00 [pmc-release]
PST - ppublish
SO  - Biochem J. 1999 Mar 1;338 ( Pt 2)(Pt 2):375-86.