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Purification and characterisation of a water-soluble ferrochelatase from Bacillus subtilis

Hansson, Mats LU and Hederstedt, Lars LU (1994) In European Journal of Biochemistry 220(1). p.201-208
Abstract
Bacillus subtilis ferrochelatase is encoded by the hemH gene of the hemEHY gene cluster and catalyses the incorporation of Fe2+ into protoporphyrin IX. B. subtilis ferrochelatase produced in Escherichia coli was purified. It was found to be a monomeric, water-soluble enzyme of molecular mass 35 kDa which in addition to Fe2+ can incorporate Zn2+ and Cu2+ into protoporphyrin IX. Chemical modification experiments indicated that the single cysteine residue in the ferrochelatase is required for enzyme activity although it is not a conserved residue compared to other ferrochelatases. In growing B. subtilis, the ferrochelatase constitutes approximately 0.05% (by mass) of the total cell protein, which corresponds to some 600 ferrochelatase... (More)
Bacillus subtilis ferrochelatase is encoded by the hemH gene of the hemEHY gene cluster and catalyses the incorporation of Fe2+ into protoporphyrin IX. B. subtilis ferrochelatase produced in Escherichia coli was purified. It was found to be a monomeric, water-soluble enzyme of molecular mass 35 kDa which in addition to Fe2+ can incorporate Zn2+ and Cu2+ into protoporphyrin IX. Chemical modification experiments indicated that the single cysteine residue in the ferrochelatase is required for enzyme activity although it is not a conserved residue compared to other ferrochelatases. In growing B. subtilis, the ferrochelatase constitutes approximately 0.05% (by mass) of the total cell protein, which corresponds to some 600 ferrochelatase molecules/cell. The turnover number of isolated ferrochelatase, 18-29 min-1, was found to be consistent with the rate of haem synthesis in exponentially growing cells (0.2 mol haem formed/min/mol enzyme). It is concluded that the B. subtilis ferrochelatase has enzymic properties which are similar to those of other characterised ferrochelatases of known primary structure, i.e. ferrochelatases of the mitochondrial inner membrane of yeast and mammalian cells. However, in contrast to these enzymes the B. subtilis enzyme is a water-soluble protein and should be more amenable to structural analysis. (Less)
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organization
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type
Contribution to journal
publication status
published
subject
keywords
Gene Deletion, Ferrochelatase/genetics/*isolation & purification/metabolism, Molecular, Escherichia coli/enzymology/genetics, Cloning, Catalysis, Amino Acid Sequence, Bacillus subtilis/*enzymology/genetics, Genes, Bacterial, Kinetics, Molecular Sequence Data, Molecular Weight, Solubility, Water
in
European Journal of Biochemistry
volume
220
issue
1
pages
201 - 208
publisher
Wiley-Blackwell
external identifiers
  • scopus:0028147501
ISSN
0014-2956
DOI
10.1111/j.1432-1033.1994.tb18615.x
language
English
LU publication?
yes
id
7ea39097-fe61-4f4f-9d10-479c1e1c40e5 (old id 8001509)
alternative location
http://www.ncbi.nlm.nih.gov/pubmed/8119288
date added to LUP
2015-10-01 09:22:51
date last changed
2017-08-14 16:38:15
@article{7ea39097-fe61-4f4f-9d10-479c1e1c40e5,
  abstract     = {Bacillus subtilis ferrochelatase is encoded by the hemH gene of the hemEHY gene cluster and catalyses the incorporation of Fe2+ into protoporphyrin IX. B. subtilis ferrochelatase produced in Escherichia coli was purified. It was found to be a monomeric, water-soluble enzyme of molecular mass 35 kDa which in addition to Fe2+ can incorporate Zn2+ and Cu2+ into protoporphyrin IX. Chemical modification experiments indicated that the single cysteine residue in the ferrochelatase is required for enzyme activity although it is not a conserved residue compared to other ferrochelatases. In growing B. subtilis, the ferrochelatase constitutes approximately 0.05% (by mass) of the total cell protein, which corresponds to some 600 ferrochelatase molecules/cell. The turnover number of isolated ferrochelatase, 18-29 min-1, was found to be consistent with the rate of haem synthesis in exponentially growing cells (0.2 mol haem formed/min/mol enzyme). It is concluded that the B. subtilis ferrochelatase has enzymic properties which are similar to those of other characterised ferrochelatases of known primary structure, i.e. ferrochelatases of the mitochondrial inner membrane of yeast and mammalian cells. However, in contrast to these enzymes the B. subtilis enzyme is a water-soluble protein and should be more amenable to structural analysis.},
  author       = {Hansson, Mats and Hederstedt, Lars},
  issn         = {0014-2956},
  keyword      = {Gene Deletion,Ferrochelatase/genetics/*isolation & purification/metabolism,Molecular,Escherichia coli/enzymology/genetics,Cloning,Catalysis,Amino Acid Sequence,Bacillus subtilis/*enzymology/genetics,Genes,Bacterial,Kinetics,Molecular Sequence Data,Molecular Weight,Solubility,Water},
  language     = {eng},
  number       = {1},
  pages        = {201--208},
  publisher    = {Wiley-Blackwell},
  series       = {European Journal of Biochemistry},
  title        = {Purification and characterisation of a water-soluble ferrochelatase from <em>Bacillus subtilis</em>},
  url          = {http://dx.doi.org/10.1111/j.1432-1033.1994.tb18615.x},
  volume       = {220},
  year         = {1994},
}