The active site and substrates binding mode of malonyl-CoA synthetase determined by transferred nuclear Overhauser effect spectroscopy, site- directed mutagenesis, and comparative modeling studies

Jin Won Jung, Jae Hyung An, Kyu Bong Na, Yu Sam Kim, Weon Tae Lee

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

The active sites and substrate bindings of Rhizobium trifolii molonyl- CoA synthetase (MCS) catalyzing the malonyl-CoA formation from malonate and CoA have been determined based on NMR spectroscopy, site-directed mutagenesis, and comparative modeling methods. The MCS-bound conformation of malonyl-CoA was determined from two-dimensional-transferred nuclear Overhauser effect spectroscopy data. MCS protein folds into two structural domains and consists of 16 α-helices, 24-β-strands, and several long loops. The core active site was determined as a wide cleft close to the end of the small C-terminal domain. The catalytic substrate malonate is placed between ATP and His206 in the MCS enzyme, supporting His206 in its catalytic role as it generates reaction intermediate, malonyl-AMP. These findings are strongly supported by previous biochemical data, as well as by the site-directed mutagenesis data reported here. This structure reveals the biochemical role as well as the substrate specificity that conservative residues of adenylate- forming enzymes have.

Original languageEnglish
Pages (from-to)1294-1303
Number of pages10
JournalProtein Science
Volume9
Issue number7
DOIs
Publication statusPublished - 2000 Jan 1

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Mutagenesis
Coenzyme A
Site-Directed Mutagenesis
Catalytic Domain
Spectrum Analysis
Ligases
Spectroscopy
Substrates
Malonyl Coenzyme A
Reaction intermediates
Rhizobium
Enzymes
Adenosine Monophosphate
Substrate Specificity
Nuclear magnetic resonance spectroscopy
Conformations
Magnetic Resonance Spectroscopy
Adenosine Triphosphate
malonyl-CoA synthetase
Proteins

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology

Cite this

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abstract = "The active sites and substrate bindings of Rhizobium trifolii molonyl- CoA synthetase (MCS) catalyzing the malonyl-CoA formation from malonate and CoA have been determined based on NMR spectroscopy, site-directed mutagenesis, and comparative modeling methods. The MCS-bound conformation of malonyl-CoA was determined from two-dimensional-transferred nuclear Overhauser effect spectroscopy data. MCS protein folds into two structural domains and consists of 16 α-helices, 24-β-strands, and several long loops. The core active site was determined as a wide cleft close to the end of the small C-terminal domain. The catalytic substrate malonate is placed between ATP and His206 in the MCS enzyme, supporting His206 in its catalytic role as it generates reaction intermediate, malonyl-AMP. These findings are strongly supported by previous biochemical data, as well as by the site-directed mutagenesis data reported here. This structure reveals the biochemical role as well as the substrate specificity that conservative residues of adenylate- forming enzymes have.",
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The active site and substrates binding mode of malonyl-CoA synthetase determined by transferred nuclear Overhauser effect spectroscopy, site- directed mutagenesis, and comparative modeling studies. / Jung, Jin Won; An, Jae Hyung; Na, Kyu Bong; Kim, Yu Sam; Lee, Weon Tae.

In: Protein Science, Vol. 9, No. 7, 01.01.2000, p. 1294-1303.

Research output: Contribution to journalArticle

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