TY - JOUR
T1 - Characterization of a novel gelatin-binding 21 kDa protein secreted by cultured adherent cells
AU - Keski-Oja, Jorma
AU - Laiho, Marikki
AU - Vartio, Tapio
N1 - Funding Information:
We thank Ms. Marja Valasjarvi and Ms. Tuija J~irvinen for fine assistance. This study was supported by the Nordisk Insulinfond, The Medical Research Council of the Academy of Finland, The Sigrid Juselius Foundation, and the Finnish Cancer Foundation.
Copyright:
Copyright 2014 Elsevier B.V., All rights reserved.
PY - 1986/7/16
Y1 - 1986/7/16
N2 - A novel gelatin-binding 21 kDa protein was identified in the culture medium of fibroblastic and sarcoma cells by affinity chromatography on gelatin-Sepharose. Its affinity for gelatin was lower than that of the other gelatin-binding proteins, fibronectin and the 70 kDa protein, as judged by stepwise elution by urea and arginine. The protein bound also to spermine and to some extent to heparin but not to staphylococcal protein A, bovine serum albumin, concanavalin A or plain Sepharose 4B. In gel filtration chromatography the protein eluted in fractions differing from those of fibronectin and the Mr 70 000 protein and retained its ability to bind to gelatin-Sepharose, indicating that the binding was not mediated by the two other gelatin-binding proteins. It contains intrachain disulfide bridges, as judged by analysis under nonreducing and reducing conditions. The protein is composed of two major subtypes with pI values of 5.85-6.10 and 6.55-6.75. It was sensitive to trypsin but not to collagenase or thrombin. Antiserum was raised in rabbits against the gelatin-binding proteins isolated from serum-free conditioned fibroblast culture medium. The antiserum reacted with fibronectin, the Mr 70 000 protein and the Mr 21 000 protein in immunoprecipitation experiments. Absorption of the antiserum with human plasma fibronectin did not decrease its reactivity with the Mr 70 000 and 21 000 proteins. However, absorption with the Mr 70 000 protein abolished also the reactivity against the Mr 21 000 protein, suggesting immunological cross-reactivity. The protein was synthesized independently from the Mr 70 000 protein, as shown by pulse-chase labeling experiments of cells. The production of the Mr 21 000 protein in cultured cells was enhanced by transforming growth factor-β.
AB - A novel gelatin-binding 21 kDa protein was identified in the culture medium of fibroblastic and sarcoma cells by affinity chromatography on gelatin-Sepharose. Its affinity for gelatin was lower than that of the other gelatin-binding proteins, fibronectin and the 70 kDa protein, as judged by stepwise elution by urea and arginine. The protein bound also to spermine and to some extent to heparin but not to staphylococcal protein A, bovine serum albumin, concanavalin A or plain Sepharose 4B. In gel filtration chromatography the protein eluted in fractions differing from those of fibronectin and the Mr 70 000 protein and retained its ability to bind to gelatin-Sepharose, indicating that the binding was not mediated by the two other gelatin-binding proteins. It contains intrachain disulfide bridges, as judged by analysis under nonreducing and reducing conditions. The protein is composed of two major subtypes with pI values of 5.85-6.10 and 6.55-6.75. It was sensitive to trypsin but not to collagenase or thrombin. Antiserum was raised in rabbits against the gelatin-binding proteins isolated from serum-free conditioned fibroblast culture medium. The antiserum reacted with fibronectin, the Mr 70 000 protein and the Mr 21 000 protein in immunoprecipitation experiments. Absorption of the antiserum with human plasma fibronectin did not decrease its reactivity with the Mr 70 000 and 21 000 proteins. However, absorption with the Mr 70 000 protein abolished also the reactivity against the Mr 21 000 protein, suggesting immunological cross-reactivity. The protein was synthesized independently from the Mr 70 000 protein, as shown by pulse-chase labeling experiments of cells. The production of the Mr 21 000 protein in cultured cells was enhanced by transforming growth factor-β.
KW - (Fibroblast)
KW - 21 kDa protein
KW - Collagen
KW - Fibronectin
KW - Gelatin binding
KW - Pericellular matrix
UR - http://www.scopus.com/inward/record.url?scp=0022497746&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0022497746&partnerID=8YFLogxK
U2 - 10.1016/0304-4165(86)90260-6
DO - 10.1016/0304-4165(86)90260-6
M3 - Article
C2 - 3015229
AN - SCOPUS:0022497746
SN - 0304-4165
VL - 882
SP - 367
EP - 376
JO - BBA - General Subjects
JF - BBA - General Subjects
IS - 3
ER -