Manipulation of prenyl chain length determination mechanism of cis-prenyltransferases

被引:49
作者
Kharel, Y [1 ]
Takahashi, S [1 ]
Yamashita, S [1 ]
Koyama, T [1 ]
机构
[1] Tohoku Univ, Inst Multidisciplinary Res Adv Mat, Aoba Ku, Sendai, Miyagi, Japan
关键词
chain-length determination mechanism; cis-prenyltransferase; isoprenoid biosynthesis; Micrococcus futeus B-P 26; undecaprenyl diphosphate synthase;
D O I
10.1111/j.1742-4658.2005.05097.x
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The carbon backbones of Z,E-mixed isoprenoids are synthesized by sequential cis-condensation of isopentenyl diphosphate (IPP) and an allylic diphosphate through actions of a series of enzymes called cis-prenyl transferases. Recent molecular analyses of Micrococcus luteus B-P 26 undecaprenyl diphosphate (UPP, C-55) synthase [Fujihashi M, Zhang Y-W, Higuchi Y, Li X-Y, Koyama T & Miki K (2001) Proc Natl Acad Sci USA 98, 4337-4342.] showed that not only the primary structure but also the crystal structure of cis-prenyltransferases were totally different from those of trans-prenyltransferases. Although many studies on structure-function relationships of cis-prenyltransferases have been reported, regulation mechanisms for the ultimate prenyl chain length have not yet been elucidated. We report here that the ultimate chain length of prenyl products can be controlled through Structural manipulation of UPP synthase of M. luteus B-P 26, based on comparisons between structures of various cis-prenyltransferases. Replacements of Ala72, Phe73, and Trp78, which are located in the proximity Of the Substrate binding site, with Leu - as in Z,E-farnesyl diphosphate (C-15) synthase - resulted in shorter ultimate products with C20-35. Additional mutation of F223H resulted in even shorter products. On the other hand, insertion of charged residues originating from long-chain cis-prenyltransferases into helix-3, which participates in constitution of the large hydrophobic cleft, resulted in lengthening of the ultimate product chain length, leading to C60-75. These results helped us understand reaction mechanisms of cis-prenyltransferase including regulation of the ultimate prenyl chain-length.
引用
收藏
页码:647 / 657
页数:11
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