Kinetic properties of spermine synthase from bovine brain.
A kinetic analysis including initial-velocity and product-inhibition studies were performed with spermine synthase purified from bovine brain. The enzyme activity was assayed in the presence of 5'-methylthioadenosine phosphorylase as an auxiliary enzyme to prevent the accumulation of the inhibitory product, 5'-methylthioadenosine, and thus to obtain linearity of the reaction with time. Initial-velocity studies gave intersecting or converging linear double-reciprocal plots. No substrate inhibition by decarboxylated S-adenosylmethionine was observed at concentrations up to 0.4 mM. Apparent Michaelis constants were 60 microM for spermidine and 0.1 microM for decarboxylated S-adenosylmethionine. Spermine was a competitive product inhibitor with respect to decarboxylated S-adenosylmethionine, but a mixed one with respect to the other substrate, spermidine. 5'-Methylthioadenosine showed a mixed inhibition with both substrates, predominantly competitive with respect to decarboxylated S-adenosylmethionine and predominantly uncompetitive with respect to spermidine. The observed kinetic and inhibition patterns are consistent with a compulsory-order mechanism, where both substrates add to the enzyme before products can be released.
D000241:Adenosine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001921:Brain / Q000201:enzymology*; D002417:Cattle; D003839:Deoxyadenosines; D007700:Kinetics; D010920:Placenta / Q000201:enzymology; D011683:Purine-Nucleoside Phosphorylase / Q000302:isolation & purification / Q000494:pharmacology; D012710:Serum Albumin, Bovine / Q000494:pharmacology; D013095:Spermidine / Q000378:metabolism; D013096:Spermine / Q000494:pharmacology; D013097:Spermine Synthase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013872:Thionucleosides / Q000494:pharmacology; D014166:Transferases / Q000378:metabolism*
D003839:Deoxyadenosines; D013872:Thionucleosides; D012710:Serum Albumin, Bovine; D013096:Spermine; C008500:5'-methylthioadenosine; D014166:Transferases; D011683:Purine-Nucleoside Phosphorylase; C021784:5'-methylthioadenosine phosphorylase; D013097:Spermine Synthase; D000241:Adenosine; D013095:Spermidine