Stassi D L, Kakavas S J, Reynolds K A, Gunawardana G, Swanson S, Zeidner D, Jackson M, Liu H, Buko A, Katz L
Pharmaceutical Products Division, Abbott Laboratories, Abbott Park, IL 60064, USA.
Proc Natl Acad Sci U S A. 1998 Jun 23;95(13):7305-9. doi: 10.1073/pnas.95.13.7305.
A previously unknown chemical structure, 6-desmethyl-6-ethylerythromycin A (6-ethylErA), was produced through directed genetic manipulation of the erythromycin (Er)-producing organism Saccharopolyspora erythraea. In an attempt to replace the methyl side chain at the C-6 position of the Er polyketide backbone with an ethyl moiety, the methylmalonate-specific acyltransferase (AT) domain of the Er polyketide synthase was replaced with an ethylmalonate-specific AT domain from the polyketide synthase involved in the synthesis of the 16-member macrolide niddamycin. The genetically altered strain was found to produce ErA, however, and not the ethyl-substituted derivative. When the strain was provided with precursors of ethylmalonate, a small quantity of a macrolide with the mass of 6-ethylErA was produced in addition to ErA. Because substrate for the heterologous AT seemed to be limiting, crotonyl-CoA reductase, a primary metabolic enzyme involved in butyryl-CoA production in streptomycetes, was expressed in the strain. The primary macrolide produced by the reengineered strain was 6-ethylErA.
通过对产生红霉素（Er）的生物红色糖多孢菌进行定向基因操作，产生了一种以前未知的化学结构，即6-去甲基-6-乙基红霉素A（6-乙基ErA）。为了用乙基部分取代Er聚酮骨架C-6位的甲基侧链，将Er聚酮合酶的丙二酸单酰特异性酰基转移酶（AT）结构域替换为参与16元大环内酯类抗生素尼达霉素合成的聚酮合酶的丙二酸二乙酯特异性AT结构域。然而，发现基因改造后的菌株产生的是ErA，而不是乙基取代的衍生物。当向该菌株提供丙二酸二乙酯的前体时，除了ErA外，还产生了少量质量为6-乙基ErA的大环内酯类抗生素。由于异源AT的底物似乎有限，因此在该菌株中表达了巴豆酰辅酶A还原酶，这是一种参与链霉菌中丁酰辅酶A产生的初级代谢酶。基因工程改造后的菌株产生的主要大环内酯类抗生素是6-乙基ErA。
