tailieunhanh - Báo cáo khoa học: Interactions between coenzyme B12 analogs and adenosylcobalamin-dependent glutamate mutase from Clostridium tetanomorphum

Adenosylcobalamin (AdoCbl)-dependent glutamate mutase fromClostrid-ium tetanomorphum comprises two weakly-associating subunits, MutS and MutE, which combine with AdoCbl to form the active holo-enzyme. Three coenzyme analogs, methylcobinamide (MeCbi), adenosylcobinamide (Ado-Cbi) and adeosylcobinamide-GDP (AdoCbi-GDP), were synthesized at milligram scale. | ễFEBS Journal Interactions between coenzyme B12 analogs and adenosylcobalamin-dependent glutamate mutase from Clostridium tetanomorphum Hao-Ping Chen1 Huei-Ju Hsu1 Fang-Ciao Hsu1 Chien-Chen Lai2 and Chung-Hua Hsu3 1 Institute of Biotechnology NationalTaipei University of Technology Taiwan 2 Institute of Molecular Biology NationalChung-Hsing University Taichung Taiwan 3 Department of AgriculturalChemistry NationalTaiwan University Taipei Taiwan Keywords adenosylcobalamin adenosylcobinamide AdoCbi-GDP B12 glutamate mutase Correspondence . Chen Institute of Biotechnology NationalTaipei University of Technology 1 Sec 3 Chung-Hsiao East Road Taipei 106 Taiwan Fax 886 2 27317117 Tel 886 2 27712171 ext. 2528 E-mail hpchen@ Received 14 August 2008 revised 30 September 2008 accepted 2 October 2008 doi Adenosylcobalamin AdoCbl -dependent glutamate mutase from Clostridium tetanomorphum comprises two weakly-associating subunits MutS and MutE which combine with AdoCbl to form the active holo-enzyme. Three coenzyme analogs methylcobinamide MeCbi adenosylcobinamide Ado-Cbi and adeosylcobinamide-GDP AdoCbi-GDP were synthesized at milligram scale. Equilibrium dialysis was used to measure the binding of coenzyme B12 analogs to glutamate mutase. Our results show that unlike AdoCbl-dependent methylmalonyl CoA mutase the ratio kcat Km decreased approximately 104-fold in both cases when AdoCbi or AdoCbi-GDP was used as the cofactor. The coenzyme analog-binding studies show that in the absence of the ribonucleotide tail of AdoCbl the enzyme s active site cannot correctly accommodate the coenzyme analog AdoCbi. The results presented here shed some light on the cobalt-carbon cleavage mechanism of B12. Glutamate mutase from Clostridium tetanomorphum is one of a group of adenosylcobalamin AdoCbl -depen-dent mutases that catalyzes the inter-conversion of L-glutamate and threo- P-methyl-L-aspartate. It comprises two weakly-associating subunits .