tailieunhanh - Báo cáo khoa học: Active site residues governing substrate selectivity and polyketide chain length in aloesone synthase

Aloesone synthase (ALS) and chalcone synthase (CHS) are plant-specific type III poyketide synthases sharing 62% amino acid sequence identity. ALS selects acetyl-CoA as a starter and carries out six successive condensa-tions with malonyl-CoA to produce a heptaketide aloesone, whereas CHS catalyses condensations of 4-coumaroyl-CoA with three malonyl-CoAs to generate chalcone. | iFEBS Journal Active site residues governing substrate selectivity and polyketide chain length in aloesone synthase Ikuro Abe1 2 Tatsuya Watanabe1 Weiwei Lou1 and Hiroshi Noguchi1 1 Schoolof PharmaceuticalSciences and the COE21 Program University of Shizuoka Japan 2 PRESTO Japan Science and Technology Agency Japan Keywords type III polyketide synthase chalcone synthase superfamily aloesone synthase chalcone synthase engineered biosynthesis Correspondence I. Abe Schoolof PharmaceuticalSciences University of Shizuoka 52-1 Yada Shizuoka 422-8526 Japan Tel. Fax 81 54 264 5662 E-mail abei@ Received 12 October 2005 revised 7 November 2005 accepted 10 November 2005 doi Aloesone synthase ALS and chalcone synthase CHS are plant-specific type III poyketide synthases sharing 62 amino acid sequence identity. ALS selects acetyl-CoA as a starter and carries out six successive condensations with malonyl-CoA to produce a heptaketide aloesone whereas CHS catalyses condensations of 4-coumaroyl-CoA with three malonyl-CoAs to generate chalcone. In ALS CHS s Thr197 Gly256 and Ser338 the active site residues lining the initiation elongation cavity are uniquely replaced with Ala Leu and Thr respectively. A homology model predicted that the active site architecture of ALS combines a horizontally restricting G256L substitution with a downward expanding T197A replacement relative to CHS. Moreover ALS has an additional buried pocket that extends into the floor of the active site cavity. The steric modulation thus facilitates ALS to utilize the smaller acetyl-CoA starter while providing adequate volume for the additional polyketide chain extensions. In fact it was demonstrated that CHS-like point mutations at these positions A197T L256G and T338S completely abolished the heptaketide producing activity. Instead A197T mutant yielded a pentaketide 2 7-dihydroxy-5-methylchro-mone while L256G and T338S just afforded a triketide triacetic .

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