Self-malonylation is an intrinsic property of a chemically synthesized type II polyketide synthase acyl carrier protein

被引:34
作者
Arthur, CJ
Szafranska, A
Evans, SE
Findlow, SC
Burston, SG
Owen, P
Clark-Lewis, I
Simpson, TJ
Crosby, J
Crump, MP
机构
[1] Univ Bristol, Sch Chem, Bristol BS8 1TS, Avon, England
[2] Univ Southampton, Sch Biol Sci, Southampton SO16 7PX, Hants, England
[3] Univ Bristol, Sch Med Sci, Dept Biochem, Bristol BS8 1TD, Avon, England
[4] Univ British Columbia, Biomed Res Ctr, Vancouver, BC V6T 1Z3, Canada
[5] Univ British Columbia, Dept Biochem & Mol Biol, Vancouver, BC V6T 1Z3, Canada
关键词
D O I
10.1021/bi051499i
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
During polyketide biosynthesis, malonyl groups are transferred to the acyl carrier protein (ACP) component of the polyketide synthase (PKS), and it has been shown that a number of type II polyketide ACPs undergo rapid self-acylation from malonyl-CoA in the absence of a malonyl-CoA:holo-acyl carrier protein transacylase (MCAT). More recently, however, the observation of self-malonylation has been ascribed to contamination with Escherichia coli MCAT (FabD) rather than an intrinsic property of the ACP. The wild-type apo-ACP from the actinorhodin (act) PKS of Streptomyces coelicolor (synthetic apo-ACP) has therefore been synthesized using solid-state peptide methods and refolded using the GroEL/ES chaperone system from E. coli. Correct folding of the act ACP has been confirmed by circular dichroism (CD) and H-1 NMR. Synthetic apo-ACP was phosphopantetheinylated to 100% by S. coelicolor holo-acyl carrier protein synthase (ACPS), and the resultant holo-ACP underwent self-malonylation in the presence of malonyl-CoA. No malonylation of negative controls was observed, confirming that the use of ACPS and GroEL/ES did not introduce contamination with E. coli MCAT. This result proves unequivocally that self-malonylation is an inherent activity of this PKS ACP in vitro.
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页码:15414 / 15421
页数:8
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