Carboxymethylproline Synthase (CarB), an Unusual Carbon-Carbon Bond-forming Enzyme of the Crotonase Superfamily Involved in Carbapenem Biosynthesis*
Carboxymethylproline Synthase (CarB), an Unusual Carbon-Carbon Bond-forming Enzyme of the Crotonase Superfamily Involved in Carbapenem Biosynthesis*
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羧甲基脯氨酸合酶 (CarB)，一种参与碳青霉烯生物合成的巴豆酶超家族的不寻常碳-碳键形成酶*
DOI：
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发表时间：
2004
影响因子：
4.8
通讯作者：
C. Schofield
中科院分区：
文献类型：
--
作者：
M. Sleeman;C. Schofield
Carboxymethylproline synthase (CarB) catalyzes the committed step in the biosynthesis of (R)-1-carbapen-2-em-3-carboxylate, the simplest member of the carbapenem family of β-lactam antibiotics, some of which are used clinically. CarB displays sequence homology with members of the crotonase family including enoyl-CoA hydratase (crotonase) and methylmalonyl-CoA decarboxylase. The CarB reaction has been proposed to comprise condensation of acetyl coenzyme A (AcCoA) and glutamate semi-aldehyde to give (2S,5S)-carboxymethylproline ((2S,5S)-CMP). (2S,5S)-CMP is then cyclized in an ATP-driven reaction catalyzed by CarA to give a carbapenam, which is subsequently epimerized and desaturated to give a carbapenem in a CarC-mediated reaction. Here we report the purification of recombinant CarB and that it exists predominantly in a trimeric form as do other members of the crotonase family. AcCoA was not found to be a substrate for CarB. Instead malonyl-CoA was found to be a substrate, efficiently producing (2S,5S)-CMP in the presence of glutamate semi-aldehyde. In the absence of glutamate semi-aldehyde, mass spectrometric analysis indicated that CarB catalyzed the decarboxylation of malonyl-CoA to AcCoA. The reactions of CarB, CarA, and CarC were coupled in vitro demonstrating the viability of malonyl-CoA as a carbapenem precursor. CarB was also shown to accept methylmalonyl CoA as a substrate to form 6-methyl-(2S,5S)CMP, which in turn is a substrate for CarA. The implications of the results for the biosynthesis of both carbapenem-3-carboxylate and C-2/C-6-substituted carbapenems, such as thienamycin, are discussed.
影响因子：
2.9
作者：
Gerratana, B;Stapon, A;Townsend, CA
通讯作者：
Townsend, CA
影响因子：
15
作者：
Stapon, A;Li, RF;Townsend, CA
通讯作者：
Townsend, CA
影响因子：
2.9
作者：
Bahnson, BJ;Anderson, VE;Petsko, GA
通讯作者：
Petsko, GA