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Metallation of the transition-state inhibitor N-methyl mesoporphyrin by ferrochelatase: Implications for the catalytic reaction mechanism

Shipovskov, Stepan LU ; Karlberg, Tobias LU ; Fodje, Michel LU ; Hansson, Mattias LU ; Ferreira, Gloria C; Hansson, Mats LU ; Reimann, Curt LU and Al-Karadaghi, Salam LU (2005) In Journal of Molecular Biology 352(5). p.1081-1090
Abstract
Insertion of metals into various tetrapyrroles is catalysed by a group of enzymes called chelatases, e.g. nickel, cobalt, magnesium and ferrochelatase. It has been proposed that catalytic metallation includes distorting the porphyrin substrate by the enzyme towards a transition state-like geometry in which at least one of the pyrrole rings will be available for metal chelation. Here, we present a study of metal insertion into the transition-state inhibitor of protoporphyrin IX ferrochelatase, N-methyl mesoporphyrin (N-MeMP), by time-resolved crystallography and mass spectrometry with and without the presence of ferrochelatase. The results show that metallation of N-MeMP has a very limited effect on the conformation of the residues that... (More)
Insertion of metals into various tetrapyrroles is catalysed by a group of enzymes called chelatases, e.g. nickel, cobalt, magnesium and ferrochelatase. It has been proposed that catalytic metallation includes distorting the porphyrin substrate by the enzyme towards a transition state-like geometry in which at least one of the pyrrole rings will be available for metal chelation. Here, we present a study of metal insertion into the transition-state inhibitor of protoporphyrin IX ferrochelatase, N-methyl mesoporphyrin (N-MeMP), by time-resolved crystallography and mass spectrometry with and without the presence of ferrochelatase. The results show that metallation of N-MeMP has a very limited effect on the conformation of the residues that participate in porphyrin and metal binding. These findings support theoretical data, which indicate that product release is controlled largely by the strain created by metal insertion into the distorted porphyrin. The results suggest that, similar to noncatalytic metallation of N-MeMP, the ferrochelatase-assisted metallation depends on the ligand exchange rate for the respective metal. Moreover, ferrochelatase catalyses insertion of Cu(II) and Zn(II) into N-MeMP with a rate that is about 20 times faster than non-enzymatic metallation in solution, suggesting that the catalytic strategy of ferrochelatase includes a stage of acceleration of the rate of ligand exchange for the metal substrate. The greater efficiency of N-MeMP metallation by Cu(II), as compared to Zn(II), contrasts with the Km values for Zn(II), (17 mu M) and Cu(II) (170 mu M) obtained for metallation of protoporphyrin IX. We suggest that this difference in metal specificity depends on the type of distortion imposed by the enzyme on protoporphyrin IX, which is different from the intrinsic non-planar distortion of N-MeMP. A mechanism of control of metal specificity by porphyrin distortion may be general for different chelatases, and may have common features with the mechanism of metal specificity in crown ethers.(c) 2005 Elsevier Ltd. All rights reserved. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Journal of Molecular Biology
volume
352
issue
5
pages
1081 - 1090
publisher
Elsevier
external identifiers
  • pmid:16140324
  • wos:000232187100007
  • scopus:24944548315
ISSN
1089-8638
DOI
10.1016/j.jmb.2005.08.002
language
English
LU publication?
yes
id
ed0e3218-18e3-4cb6-a8ce-681bc239b166 (old id 150932)
date added to LUP
2007-06-27 15:41:37
date last changed
2017-02-19 04:18:21
@article{ed0e3218-18e3-4cb6-a8ce-681bc239b166,
  abstract     = {Insertion of metals into various tetrapyrroles is catalysed by a group of enzymes called chelatases, e.g. nickel, cobalt, magnesium and ferrochelatase. It has been proposed that catalytic metallation includes distorting the porphyrin substrate by the enzyme towards a transition state-like geometry in which at least one of the pyrrole rings will be available for metal chelation. Here, we present a study of metal insertion into the transition-state inhibitor of protoporphyrin IX ferrochelatase, N-methyl mesoporphyrin (N-MeMP), by time-resolved crystallography and mass spectrometry with and without the presence of ferrochelatase. The results show that metallation of N-MeMP has a very limited effect on the conformation of the residues that participate in porphyrin and metal binding. These findings support theoretical data, which indicate that product release is controlled largely by the strain created by metal insertion into the distorted porphyrin. The results suggest that, similar to noncatalytic metallation of N-MeMP, the ferrochelatase-assisted metallation depends on the ligand exchange rate for the respective metal. Moreover, ferrochelatase catalyses insertion of Cu(II) and Zn(II) into N-MeMP with a rate that is about 20 times faster than non-enzymatic metallation in solution, suggesting that the catalytic strategy of ferrochelatase includes a stage of acceleration of the rate of ligand exchange for the metal substrate. The greater efficiency of N-MeMP metallation by Cu(II), as compared to Zn(II), contrasts with the Km values for Zn(II), (17 mu M) and Cu(II) (170 mu M) obtained for metallation of protoporphyrin IX. We suggest that this difference in metal specificity depends on the type of distortion imposed by the enzyme on protoporphyrin IX, which is different from the intrinsic non-planar distortion of N-MeMP. A mechanism of control of metal specificity by porphyrin distortion may be general for different chelatases, and may have common features with the mechanism of metal specificity in crown ethers.(c) 2005 Elsevier Ltd. All rights reserved.},
  author       = {Shipovskov, Stepan and Karlberg, Tobias and Fodje, Michel and Hansson, Mattias and Ferreira, Gloria C and Hansson, Mats and Reimann, Curt and Al-Karadaghi, Salam},
  issn         = {1089-8638},
  language     = {eng},
  number       = {5},
  pages        = {1081--1090},
  publisher    = {Elsevier},
  series       = {Journal of Molecular Biology},
  title        = {Metallation of the transition-state inhibitor N-methyl mesoporphyrin by ferrochelatase: Implications for the catalytic reaction mechanism},
  url          = {http://dx.doi.org/10.1016/j.jmb.2005.08.002},
  volume       = {352},
  year         = {2005},
}