Characterization of a novel spermidine/spermine acetyltransferase, BltD, from Bacillus subtilis.
Overexpression of the BltD gene in Bacillus subtilis causes acetylation of the polyamines spermidine and spermine. BltD is co-regulated with another gene, Blt, which encodes a multidrug export protein whose overexpression facilitates spermidine export [Woolridge, Vazquez-Laslop, Markham, Chevalier, Gerner and Neyfakh (1997) J. Biol. Chem. 272, 8864-8866]. Here we show that BltD acetylates both spermidine and spermine at primary propyl amine moieties, with spermine being the preferred substrate. In the presence of saturating concentrations of acetyl CoA, BltD rapidly acetylates spermine at both the N1 and N12 positions. The Km (app) values for spermine, spermidine and N1-acetylspermine are </=67, 200 and 1200 microM, respectively. Diamines ranging from 1, 3-diaminopropane to 1,12-diaminododecane, monoacetylputrescine and N8-acetylspermidine were not substrates for BltD. Putrescine (1, 4-diaminobutane) and N8-acetylspermidine were competitive inhibitors of spermidine acetylation by BltD, with Ki values of 0.25 and 5.76 mM, respectively. CoA competitively inhibited both spermidine and acetyl-CoA interactions with BltD. These data and other results indicate that the mechanism of spermidine and spermine acetylation by BltD is a random-order mechanism of bi-molecular kinetics.
Woolridge
D P
DP
Department of Biochemistry, University of Arizona, Arizona Cancer Center, 1515 N. Campbell Avenue, Tucson, AZ 85724, USA.
Martinez
J D
JD
Stringer
D E
DE
Gerner
E W
EW
D000105:Acetyl Coenzyme A / Q000378:metabolism; D000107:Acetylation; D000123:Acetyltransferases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D001412:Bacillus subtilis / Q000201:enzymology*; D001667:Binding, Competitive; D003065:Coenzyme A / Q000378:metabolism; D003959:Diamines / Q000378:metabolism; D007700:Kinetics; D013095:Spermidine / Q000031:analogs & derivatives / Q000378:metabolism; D013096:Spermine / Q000031:analogs & derivatives / Q000378:metabolism; D013379:Substrate Specificity; D013816:Thermodynamics; D000099228:Diamine N-Acetyltransferase
D000105:Acetyl Coenzyme A; D000123:Acetyltransferases; D003065:Coenzyme A; D003959:Diamines; D013095:Spermidine; D013096:Spermine; D000099228:Diamine N-Acetyltransferase