Functional analysis of the human tissue-type plasminogen activator protein: the light chain.
A pBR322::Rous sarcoma virus(RSV)-based shuttle vector was used to insert fused genes, composed of the amino-terminal portion of the bacterial chloramphenicol-acetyltransferase gene (cat) and the entire coding region for the C-terminally derived light (L) chain of human tissue-type plasminogen activator (t-PA) cDNA. Cotransfection of rat 3Y1 cells with pRSVneo DNA and pRSVcat/t-PA DNA yielded stably integrated G418-resistant transfectants which contain unrearranged copies of pRSVcat/t-PA DNA. These transfectants synthesize cat/t-PA L-chain mRNA, apparently correctly initiated and terminated. With the help of an enzyme-linked immunosorbent assay (ELISA), it is demonstrated that these cells produce human t-PA antigen. Furthermore, pRSVcat/t-PA L-chain cDNA-containing rat 3Y1 cells synthesize a plasminogen-dependent amidolytic activity which is suppressed by specific anti-human t-PA antibodies. This activity cannot be stimulated by fibrin, a property displayed by native t-PA. It is concluded that the t-PA L-chain cDNA contains the complete genetic information for the plasminogen activator activity.
D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics; D010450:Endopeptidases / Q000378:metabolism; D004789:Enzyme Activation; D005337:Fibrin / Q000378:metabolism; D005822:Genetic Vectors; D006801:Humans; D046911:Macromolecular Substances; D051381:Rats; D012697:Serine Endopeptidases; D013329:Structure-Activity Relationship; D013379:Substrate Specificity; D010959:Tissue Plasminogen Activator / Q000235:genetics / Q000276:immunology / Q000502:physiology*
D046911:Macromolecular Substances; D005337:Fibrin; D004247:DNA; D010450:Endopeptidases; D012697:Serine Endopeptidases; D010959:Tissue Plasminogen Activator