Magnesium protoporphyrin IX methyltransferase (ChlM), catalyses the methylation of magnesium protoporphyrin IX (MgP) at the C6 propionate side chain to form magnesium protoporphyrin IX monomethylester (MgPME). Threading methods biased by sequence similarity and predicted secondary structure have been used to assign this enzyme to a particular class of S-adenosyl-l-methionine (SAM)-binding proteins. These searches suggest that ChlM contains a seven-stranded β-sheet, common among small-molecule methyltransferases. Steady-state kinetic assays were performed using magnesium deuteroporphyrin IX (MgD), a more water-soluble substrate analogue of MgP. Initial rate studies showed that the reaction proceeds via a ternary complex. Product (S-adenosyl-l-homocysteine; SAH) inhibition was used to investigate the kinetic mechanism further. SAH was shown to exhibit competitive inhibition with respect to SAM, and mixed inhibition with respect to MgD. This is indicative of a random binding mechanism, whereby SAH may bind productively to either free enzyme or a ChlM–MgD complex. Our results provide an overview of the steady-state kinetics for this enzyme, which are significant given the role of MgP and MgPME in plastid-to-nucleus signalling and their likely critical role in the regulation of this biosynthetic pathway.
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April 2003
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Research Article|
April 15 2003
Purification and kinetic characterization of the magnesium protoporphyrin IX methyltransferase from Synechocystis PCC6803
Mark SHEPHERD;
Mark SHEPHERD
Robert Hill Institute for Photosynthesis, Department of Molecular Biology and Biotechnology, Firth Court, Western Bank, University of Sheffield, Sheffield S10 2TN, U.K.
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James D. REID;
James D. REID
Robert Hill Institute for Photosynthesis, Department of Molecular Biology and Biotechnology, Firth Court, Western Bank, University of Sheffield, Sheffield S10 2TN, U.K.
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C. Neil HUNTER
C. Neil HUNTER
1
Robert Hill Institute for Photosynthesis, Department of Molecular Biology and Biotechnology, Firth Court, Western Bank, University of Sheffield, Sheffield S10 2TN, U.K.
1To whom correspondence should be addressed (e-mail C.N.Hunter@sheffield.ac.uk).
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Publisher: Portland Press Ltd
Received:
September 04 2002
Revision Received:
November 21 2002
Accepted:
December 18 2002
Accepted Manuscript online:
December 18 2002
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London ©2003
2003
Biochem J (2003) 371 (2): 351–360.
Article history
Received:
September 04 2002
Revision Received:
November 21 2002
Accepted:
December 18 2002
Accepted Manuscript online:
December 18 2002
Citation
Mark SHEPHERD, James D. REID, C. Neil HUNTER; Purification and kinetic characterization of the magnesium protoporphyrin IX methyltransferase from Synechocystis PCC6803. Biochem J 15 April 2003; 371 (2): 351–360. doi: https://doi.org/10.1042/bj20021394
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