Post‐translational modification of heterologously expressed Streptomyces type II polyketide synthase acyl carrier proteins

FEBS Letters

Tanner, Julian A¹  Cox, Russell J¹  Crosby, John¹  Simpson, Thomas J¹  Byrom, Kate J¹  Hitchman, Timothy S¹  Findlow, I.Stuart C¹ 

[1] School of Chemistry (University of Bristol Molecular Recognition Centre), University of Bristol, Cantock's Close, Bristol BS8 1TS, UK

关键词: Polyketide;    Acyl carrier protein;    Streptomyces;    Heterologous expression;    Post-translational modification;    4′-Phosphopantetheine;   

DOI  :  10.1016/S0014-5793(97)00202-0

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Expression in Escherichia coli of Streptomyces acyl carrier proteins (ACPs) associated with polyketide biosynthesis using the pT7-7 expression system of Tabor and Richardson led to the production predominantly of inactive apo-proteins lacking the 4′-phosphopantetheinyl prosthetic group essential for polyketide synthase activity. Modification of growth conditions led to an increase of production of active holo-protein for the actinorhodin (act) ACP, but this technique was ineffective for oxytetracycline (otc) and griseusin (gris) ACPs. Labelling experiments revealed that a low level of otc ACP expressed prior to induction was produced mainly as active holo-protein, while post-induction ¹⁵N-labelled protein was almost exclusively in the apo-ACP form. Limiting endogenous holo-acyl carrier protein synthase (ACPS) concentration was implicated as responsible for low apo-ACP to holo-ACP conversion, rather than limiting substrate (coenzyme A) and cofactor (Mg²⁺) concentrations. Co-expression of act and gris ACPs with ACPS in E. coli led to high levels of production of active holo-ACPs and ACPS. We have also made the significant observation that ACPS is able to transfer acylated CoA moieties to act apo-ACP.

【 授权许可】

Unknown