Disruption of interkringle disulfide bond of plasminogen kringle 1-3 changes the lysine binding capability of kringle 2, but not its antiangiogenic activity.
Kringle 1-3 of human plasminogen is a potent inhibitor of endothelial cell proliferation. To understand a possible role for the unique cystine bridge between kringle 2 and kringle 3, we disrupted the interkringle disulfide bond by mutating Cys(169) and Cys(297) to serine residues. The yield of the mutant during the refolding process was decreased significantly. Anti-endothelial cell proliferative activity of the mutant was similar to that of the wild type. There was no significant difference in in vivo antiangiogenic activity between the wild type and the mutant in chorioallantoic membrane assay. However, in the mutant, the weak lysine binding capability of kringle 2 was not detected and its mobility in nonreducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis is different from that of the wild type. These results support the notion that the overall antiangiogenic function of angiostatin is mediated by individual kringles, and suggest that the lysine binding capability of kringle 2 is likely not important for the antiangiogenic activity of kringle 1-3.
Lee
H
H
Mogam Biotechnology Research Institute, Yongin, 449-910, Korea.
Kim
H K
HK
Lee
J H
JH
You
W K
WK
Chung
S I
SI
Chang
S I
SI
Park
M H
MH
Hong
Y K
YK
Joe
Y A
YA
D019943:Amino Acid Substitution / Q000235:genetics; D020533:Angiogenesis Inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D002417:Cattle; D002455:Cell Division / Q000187:drug effects; D002642:Chick Embryo; D002823:Chorion / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D004220:Disulfides / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D006801:Humans; D018082:Kringles / Q000235:genetics / Q000502:physiology*; D008024:Ligands; D008239:Lysine / Q000378:metabolism*; D009154:Mutation / Q000235:genetics; D018919:Neovascularization, Physiologic / Q000187:drug effects; D010958:Plasminogen / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D011485:Protein Binding; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology; D013816:Thermodynamics
D020533:Angiogenesis Inhibitors; D004220:Disulfides; D008024:Ligands; D011994:Recombinant Proteins; D010958:Plasminogen; D008239:Lysine; D003545:Cysteine