Structural and Biochemical Characterization of ZhuI Aromatase/Cyclase from the R1128 Polyketide Pathway

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• 作者：Brian D. Ames ; Ming-Yue Lee ; Colleen Moody ; Wenjun Zhang ; Yi Tang ; Shiou-Chuan Tsai
• 刊名：Biochemistry
• 出版年：2011
• 出版时间：October 4, 2011
• 年：2011
• 卷：50
• 期：39
• 页码：8392-8406
• 全文大小：903K
• 年卷期：v.50,no.39(October 4, 2011)
• ISSN：1520-4995

文摘

Aromatic polyketides comprise an important class of natural products that possess a wide range of biological activities. The cyclization of the polyketide chain is a critical control point in the biosynthesis of aromatic polyketides. The aromatase/cyclases (ARO/CYCs) are an important component of the type II polyketide synthase (PKS) and help fold the polyketide for regiospecific cyclizations of the first ring and/or aromatization, promoting two commonly observed first-ring cyclization patterns for the bacterial type II PKSs: C7鈥揅12 and C9鈥揅14. We had previously reported the crystal structure and enzymological analyses of the TcmN ARO/CYC, which promotes C9鈥揅14 first-ring cyclization. However, how C7鈥揅12 first-ring cyclization is controlled remains unresolved. In this work, we present the 2.4 脜 crystal structure of ZhuI, a C7鈥揅12-specific first-ring ARO/CYC from the type II PKS pathway responsible for the production of the R1128 polyketides. Though ZhuI possesses a helix-grip fold shared by TcmN ARO/CYC, there are substantial differences in overall structure and pocket residue composition that may be important for directing C7鈥揅12 (rather than C9鈥揅14) cyclization. Docking studies and site-directed mutagenesis coupled to an in vitro activity assay demonstrate that ZhuI pocket residues R66, H109, and D146 are important for enzyme function. The ZhuI crystal structure helps visualize the structure and putative dehydratase function of the didomain ARO/CYCs from KR-containing type II PKSs. The sequence鈥搒tructure鈥揻unction analysis described for ZhuI elucidates the molecular mechanisms that control C7鈥揅12 first-ring polyketide cyclization and builds a foundation for future endeavors into directing cyclization patterns for engineered biosynthesis of aromatic polyketides.