Mechanism of Product Chain Length Determination and the Role of a Flexible Loop in Escherichia coli Undecaprenyl-pyrophosphate Synthase Catalysis

Tzu Ping Ko, Yi Kai Chen, Howard Robinson, Pei Chun Tsai, Yi Gui Gao, Annie P.C. Chen, Andrew H.J. Wang, Po Huang Liang

研究成果: 雜誌貢獻文章同行評審

82 引文 斯高帕斯(Scopus)

摘要

The Escherichia coli undecaprayl-pyrophosphate synthase (UPPs) structure has been solved using the single wavelength anomalous diffraction method. The putative substrate-binding site is located near the end of the βA-strand with Asp-26 playing a critical catalytic role. In both subunits, an elongated hydrophobic tunnel is found, surrounded by four β-strands (βA-βB-βD-βC) and two helices (α2 and α3) and lined at the bottom with large residues Ile-62, Leu-137, Val-105, and His-103. The product distributions formed by the use of the I62A, V105A, and H103A mutants are similar to those observed for wild-type UPPs. Catalysis by the L137A UPPs, on the other hand, results in predominantly the formation of the C70 polymer rather than the C55 polymer. Ala-69 and Ala-143 are located near the top of the tunnel. In contrast to the A143V reaction, the C30 intermediate is formed to a greater extent and is longer lived in the process catalyzed by the A69L mutant. These findings suggest that the small side chain of Ala-69 is required for rapid elongation to the C55 product, whereas the large hydrophobic side chain of Leu-137 is required to limit the elongation to the C55 product. The roles of residues located on a flexible loop were investigated. The S71A, N74A, or R77A mutants displayed 25-200-fold decrease in kcat values. W75A showed an 8-fold increase of the FPP Km value, and 22-33-fold increases in the IPP Km values were observed for E81A and S71A. The loop may function to bridge the interaction of IPP with FPP, needed to initiate the condensation reaction and serve as a hinge to control the substrate binding and product release.

原文英語
頁(從 - 到)47474-47482
頁數9
期刊Journal of Biological Chemistry
276
發行號50
DOIs
出版狀態已發佈 - 12月 14 2001
對外發佈

ASJC Scopus subject areas

  • 生物化學
  • 分子生物學
  • 細胞生物學

指紋

深入研究「Mechanism of Product Chain Length Determination and the Role of a Flexible Loop in Escherichia coli Undecaprenyl-pyrophosphate Synthase Catalysis」主題。共同形成了獨特的指紋。

引用此