Identification of a guanosine triphosphate-binding site on guinea pig liver transglutaminase. Role of GTP and calcium ions in modulating activity.
Guanosine 5'-triphosphate (GTP) was found to inhibit guinea pig liver transglutaminase activity as measured by [3H]putrescine incorporation into casein. GDP and GTP-gamma-S also inhibited enzyme activity (GTP-gamma-S greater than GTP greater than GDP). Kinetic studies showed that GTP acted as a reversible, noncompetitive inhibitor and that CaCl2 partially reversed GTP inhibition. GTP also inhibited rat liver and adult bovine aortic endothelial cell transglutaminase, but did not inhibit Factor XIIIa activity. Guanosine monophosphate (GMP), cyclic GMP, and polyguanylic acid did not inhibit enzyme activity. Guinea pig liver transglutaminase adsorbed well to GTP-agarose affinity columns, but not to CTP-agarose columns, and the binding was inhibited by the presence of calcium ions. Specific binding of GTP to transglutaminase was demonstrated by photoaffinity labeling with 8-azidoguanosine 5'-[gamma-32P] triphosphate, which was inhibited by the presence of GTP or CaCl2. GTP inhibited trypsin proteolysis of guinea pig liver transglutaminase without affecting the trypsin proteolysis of chromogenic substrates. Proteolytic protection was reversed by the addition of calcium. This study demonstrates that GTP binds to transglutaminase and that both GTP and calcium ions function in concert to regulate transglutaminase structure and function.
D000345:Affinity Labels / Q000378:metabolism; D000818:Animals; D001386:Azides / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D002122:Calcium Chloride / Q000494:pharmacology; D002846:Chromatography, Affinity; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism*; D006168:Guinea Pigs; D008099:Liver / Q000201:enzymology*; D008274:Magnesium / Q000378:metabolism; D011503:Transglutaminases / Q000378:metabolism*; D014357:Trypsin / Q000378:metabolism
C021961:8-azidoguanosine triphosphate; D000345:Affinity Labels; D001386:Azides; D006160:Guanosine Triphosphate; D011503:Transglutaminases; D014357:Trypsin; D008274:Magnesium; D002122:Calcium Chloride; D002118:Calcium