PMID:9108302
| Citation |
Cox, RJ, Hitchman, TS, Byrom, KJ, Findlow, IS, Tanner, JA, Crosby, J and Simpson, TJ (1997) Post-translational modification of heterologously expressed Streptomyces type II polyketide synthase acyl carrier proteins. FEBS Lett. 405:267-72 |
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| Abstract |
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. |
| Links | |
| Keywords |
Acyl Carrier Protein/metabolism; Apoproteins/metabolism; Coenzyme A/metabolism; Disulfides; Escherichia coli/enzymology; Multienzyme Complexes/metabolism; Protein Processing, Post-Translational; Recombinant Proteins; Species Specificity; Streptomyces/enzymology |
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