Chemical modification of alpha2-macroglobulin to generate derivatives that bind transforming growth factor-beta with increased affinity.
alpha2-Macroglobulin (alpha2M) binds a number of cytokines, including transforming growth factor-beta1 (TGF-beta1) and TGF-beta2. The affinity of these interactions depends on the alpha2M conformation. In this investigation, we treated human alpha2M with cis-dichlorodiammineplatinum (II) (cis-Pt), a crosslinking reagent that partially 'locks' the alpha2M conformation, and then with methylamine to generate a preparation (alpha2M-P/M) consisting of stable alpha2M conformational intermediates. alpha2M-P/M bound TGF-beta1 and TGF-beta2 with higher affinity than any other form of alpha2M studied to date. The equilibrium dissociation constants were 14 and 2 nM for TGF-beta1 and TGF-beta2, respectively. alpha2M-P/M, at 100 nM, neutralized the activity of TGF-beta1 by about 75% in an endothelial cell proliferation assay. The equivalent concentration of native alpha2M or methylamine-modified alpha2M had no effect. These studies demonstrate that the potential of alpha2M as a cytokine carrier and neutralizer may not be fully realized in either the native or completely activated conformations.
Webb
D J
DJ
Department of Pathology, University of Virginia Health Sciences Center, Charlottesville 22908, USA.
Gonias
S L
SL
D000818:Animals; D002478:Cells, Cultured; D002945:Cisplatin / Q000494:pharmacology; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004727:Endothelium / Q000166:cytology; D006801:Humans; D007457:Iodine Radioisotopes; D008744:Methylamines / Q000494:pharmacology; D016212:Transforming Growth Factor beta / Q000378:metabolism*; D000511:alpha-Macroglobulins / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*
D003432:Cross-Linking Reagents; D007457:Iodine Radioisotopes; D008744:Methylamines; D016212:Transforming Growth Factor beta; D000511:alpha-Macroglobulins; C027451:methylamine; D002945:Cisplatin