Pieper R, Luo G, Cane D E, Khosla C
Department of Chemical Engineering, Stanford University, California 94305-5025, USA.
Nature. 1995 Nov 16;378(6554):263-6. doi: 10.1038/378263a0.
Modular polyketide synthases (PKSs) are complex multi-enzyme proteins that catalyse the bacterial biosynthesis of many pharmaceutically useful polyketides. The PKSs are organized into a series of modules, each containing the active catalytic sites required for one step in the synthesis process. Here we report a method for cell-free enzymatic synthesis of 6-deoxyerythronolide B (6-dEB), the parent molecule of the antibiotic erythromycin A, using recombinant 6-deoxyerythronolide B synthase (DEBS), a modular PKS with at least 28 distinct active sites. We have also synthesized in vitro a triketide lactone by using a truncated mutant of DEBS. The availability of such cell-free synthetic routes will allow direct investigation of the structural and mechanistic basis for the unusual combination of high substrate specificity and tolerance to genetic reprogramming found in this enzyme family.
模块化聚酮合酶（PKSs）是复杂的多酶蛋白，可催化许多具有药学用途的聚酮化合物的细菌生物合成。PKSs被组织成一系列模块，每个模块都包含合成过程中一步所需的活性催化位点。在此，我们报告了一种使用重组6-脱氧红霉内酯B合酶（DEBS）进行无细胞酶促合成6-脱氧红霉内酯B（6-dEB）的方法，6-dEB是抗生素红霉素A的母体分子，DEBS是一种具有至少28个不同活性位点的模块化PKS。我们还使用DEBS的截短突变体在体外合成了三酮内酯。这种无细胞合成途径的可用性将允许直接研究该酶家族中高底物特异性和对基因重编程的耐受性这种异常组合的结构和机制基础。
