Dataset Information

Structural insights into the catalytic mechanism of Synechocystis magnesium protoporphyrin IX O-methyltransferase (ChlM).

ABSTRACT: Magnesium protoporphyrin IX O-methyltransferase (ChlM) catalyzes transfer of the methyl group from S-adenosylmethionine to the carboxyl group of the C13 propionate side chain of magnesium protoporphyrin IX. This reaction is the second committed step in chlorophyll biosynthesis from protoporphyrin IX. Here we report the crystal structures of ChlM from the cyanobacterium Synechocystis sp. PCC 6803 in complex with S-adenosylmethionine and S-adenosylhomocysteine at resolutions of 1.6 and 1.7 Å, respectively. The structures illustrate the molecular basis for cofactor and substrate binding and suggest that conformational changes of the two "arm" regions may modulate binding and release of substrates/products to and from the active site. Tyr-28 and His-139 were identified to play essential roles for methyl transfer reaction but are not indispensable for cofactor/substrate binding. Based on these structural and functional findings, a catalytic model is proposed.

SUBMITTER: Chen X

PROVIDER: S-EPMC4162172 | biostudies-literature | 2014 Sep

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Structural insights into the catalytic mechanism of Synechocystis magnesium protoporphyrin IX O-methyltransferase (ChlM).

Chen Xuemin X Wang Xiao X Feng Juan J Chen Yuhong Y Fang Ying Y Zhao Shun S Zhao Aiguo A Zhang Min M Liu Lin L

The Journal of biological chemistry 20140730 37

Magnesium protoporphyrin IX O-methyltransferase (ChlM) catalyzes transfer of the methyl group from S-adenosylmethionine to the carboxyl group of the C13 propionate side chain of magnesium protoporphyrin IX. This reaction is the second committed step in chlorophyll biosynthesis from protoporphyrin IX. Here we report the crystal structures of ChlM from the cyanobacterium Synechocystis sp. PCC 6803 in complex with S-adenosylmethionine and S-adenosylhomocysteine at resolutions of 1.6 and 1.7 Å, resp ...[more]

PMID: 25077963

Similar Datasets

Project description:The I, D and H subunits (ChlI, ChlD and ChlH respectively) of the magnesium protoporphyrin IX chelatase from Synechocystis have been purified to homogeneity as a result of the overexpression of the encoding genes in Escherichia coli and the production of large quantities of histidine-tagged proteins. These subunits have been used in an initial investigation of the biochemical and kinetic properties of the enzyme. The availability of pure ChlI, ChlD and ChlH has allowed us to estimate the relative concentrations of the three protein components required for optimal activity, and to investigate the dependence of chelatase activity on the concentrations of MgCl2, ATP and protoporphyrin IX. It was found that, whereas ChlD and ChlH are likely to be monomeric, ChlI can aggregate in an ATP-dependent manner, changing from a dimeric to an octameric structure. Subunit titration assays suggest an optimal ratio of ChlI, ChlD and ChlH of 2:1:4 respectively. However, the dependence of chelatase activity on increasing concentrations of ChlI and ChlH with respect to ChlD suggests that these two subunits, at least in vitro, behave as substrates in their interaction with ChlD. Mg chelation could not be detected unless the Mg2+ concentration exceeded the ATP concentration, suggesting at least two requirements for Mg2+, one as a component of MgATP2-, the other as the chelated metal. The steady-state kinetic parameters were determined from continuous assays; the Km values for protoporphyrin, MgCl2 and ATP were 1.25 microM, 4.9 mM and 0.49 mM respectively. The rate dependence of Mg2+ was clearly sigmoidal with a Hill coefficient of 3, suggesting positive co-operativity. Initiating the reaction by the addition of one of the substrates in these continuous assays resulted in a significant lag period of at least 10 min before the linear production of Mg protoporphyrin. This lag was significantly decreased by preincubating ChlI and ChlD with ATP and MgCl2, and by mixing it with ChlH that had been preincubated with protoporphyrin IX, ATP and MgCl2. This suggests not only a close MgATP2--dependent interaction between ChlI and ChlD but also an interaction between ChlH and the protoporphyrin substrate that also is stimulated by ATP and MgCl2.

Dataset Information

Structural insights into the catalytic mechanism of Synechocystis magnesium protoporphyrin IX O-methyltransferase (ChlM).

Publications

Structural insights into the catalytic mechanism of Synechocystis magnesium protoporphyrin IX O-methyltransferase (ChlM).

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets