A structural and functional study on the 2-C-methyl-d-erythritol-4-phosphate cytidyltransferase (IspD) from Bacillus subtilis.
2-C-Methyl-D-erythritol-4-phosphate cytidyltransferase (IspD) is an essential enzyme in the mevalonate-independent pathway of isoprenoid biosynthesis. This enzyme catalyzes 2-C-Methyl-d-erythritol 4-phosphate (MEP) and cytosine triphosphate (CTP) to 4-diphosphocytidyl-2-C-methyl-d-erythritol (CDPME) and inorganic pyrophosphate (PPi). Bacillus subtilis was a kind of excellent isoprene producer. However, the studies on the key enzymes of MEP pathway in B. subtilis were still absent. In this work, the crystal structures of IspD and IspD complexed with CTP from B.subtilis were determined. For the first time, the intact P-loop was observed in the apo structure of IspD enzyme. Structural comparisons revealed that the concerted movements of the P-loop and loops close to the active site were essential in the reaction catalyzed by IspD. Meanwhile, kinetic analysis showed that the CTP hydrolytic activity of IspD from B.subtilis was over two times higher than that from Escherichia coli. These results will be useful for future target-based screening of potential inhibitors and the metabolic engineering for isoprenoid biosynthesis.
Key Laboratory of Environmental and Applied Microbiology, Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu, 610041, China.|Key Laboratory of Environmental Microbiology of Sichuan Province, Chengdu, 610041, China.|University of Chinese Academy of Sciences, Beijing, 100049, China.
Yun
Y
Jin
Key Laboratory of Environmental and Applied Microbiology, Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu, 610041, China.|Key Laboratory of Environmental Microbiology of Sichuan Province, Chengdu, 610041, China.
Zhongchuan
Z
Liu
Key Laboratory of Environmental and Applied Microbiology, Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu, 610041, China.|Key Laboratory of Environmental Microbiology of Sichuan Province, Chengdu, 610041, China.
Yanjie
Y
Li
Chinese Academy of Sciences Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, People's Republic of China.
Weifeng
W
Liu
Chinese Academy of Sciences Key Laboratory of Microbial Physiological and Metabolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, People's Republic of China.
Yong
Y
Tao
Key Laboratory of Environmental and Applied Microbiology, Chengdu Institute of Biology, Chinese Academy of Sciences, Chengdu, 610041, China.|Key Laboratory of Environmental Microbiology of Sichuan Province, Chengdu, 610041, China.
Ganggang
G
Wang
D001412:Bacillus subtilis / Q000737:chemistry / Q000201:enzymology*; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D002384:Catalysis; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D003570:Cytidine Triphosphate / Q000378:metabolism*; D008958:Models, Molecular; D010088:Oxidoreductases / Q000737:chemistry* / Q000378:metabolism*; D011485:Protein Binding; D017433:Protein Structure, Secondary
D001426:Bacterial Proteins; D003570:Cytidine Triphosphate; D010088:Oxidoreductases