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Bacillus subtilis mutant succinate dehydrogenase lacking covalently bound flavin: Identification of the primary defect and studies on the iron-sulfur clusters in mutated and wild- type enzyme

Maguire, John J.; Magnusson, Kerstin and Hederstedt, Lars LU (1986) In Biochemistry 25(18). p.5202-5208
Abstract
Succinate dehydrogenase consists of two protein subunits and contains one FAD and three iron-sulfur clusters. The flavin is covalently bound to a histidine in the larger, Fp, subunit. The reduction oxidation midpoint potentials of the clusters designated S-l, S-2, and S-3 in Bacillus subtilis wild-type membrane-bound enzyme were determined as +80, -240, and -25 mV, respectively. Magnetic spin interactions between clusters S-l and S-2 and between S-l and S-3 were detected by using EPR spectroscopy. The point mutations of four B. subtilis mutants with defective Fp subunits were mapped. The gene of the mutant specifically lacking covalently bound flavin in the enzyme was cloned. The mutation was determined from the DNA sequence as a glycine... (More)
Succinate dehydrogenase consists of two protein subunits and contains one FAD and three iron-sulfur clusters. The flavin is covalently bound to a histidine in the larger, Fp, subunit. The reduction oxidation midpoint potentials of the clusters designated S-l, S-2, and S-3 in Bacillus subtilis wild-type membrane-bound enzyme were determined as +80, -240, and -25 mV, respectively. Magnetic spin interactions between clusters S-l and S-2 and between S-l and S-3 were detected by using EPR spectroscopy. The point mutations of four B. subtilis mutants with defective Fp subunits were mapped. The gene of the mutant specifically lacking covalently bound flavin in the enzyme was cloned. The mutation was determined from the DNA sequence as a glycine to aspartate substitution at a conserved site seven residues downstream from the hisitidine that binds the flavin in wild-type enzyme. The redox midpoint potential of the iron-sulfur clusters and the magnetic spin interactions in mutated succinate dehdyrogenases were indistinguishable from the those of the wild type. This shows that flavin has no role in the measured magnetic spin interactions or in the structure and stability of the iron-sulfur clusters. It is concluded from sequence and mutant studies that conserved amino acid residues around the histidyl-FAD are important for FAD binding; however, amino acids located more than 100 residues downstream from the histidyl in the Fp subunit can also effect
flavinylation. (Less)
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author
publishing date
type
Contribution to journal
publication status
published
subject
in
Biochemistry
volume
25
issue
18
pages
5202 - 5208
publisher
The American Chemical Society
ISSN
0006-2960
DOI
10.1021/bi00366a033
language
English
LU publication?
no
id
2c9d5c15-73fe-4c69-881d-483720083b6c
date added to LUP
2017-07-18 11:09:21
date last changed
2017-09-26 14:42:00
@article{2c9d5c15-73fe-4c69-881d-483720083b6c,
  abstract     = {Succinate dehydrogenase consists of two protein subunits and contains one FAD and three iron-sulfur clusters. The flavin is covalently bound to a histidine in the larger, Fp, subunit. The reduction oxidation midpoint potentials of the clusters designated S-l, S-2, and S-3 in Bacillus subtilis wild-type  membrane-bound enzyme were determined as +80, -240, and -25 mV, respectively. Magnetic spin interactions between clusters S-l and S-2 and between S-l and S-3 were detected by using EPR spectroscopy. The point mutations of four B. subtilis mutants with defective Fp subunits were mapped. The gene of the mutant specifically lacking covalently bound flavin in the enzyme was cloned. The mutation was determined from the DNA sequence as a glycine to aspartate substitution at a conserved site seven residues downstream from the hisitidine that binds the flavin in wild-type enzyme. The redox midpoint potential of the iron-sulfur clusters and the magnetic spin interactions in mutated succinate dehdyrogenases were indistinguishable from the those of the wild type. This shows that flavin has no role in the measured magnetic spin interactions or in the structure and stability of the iron-sulfur clusters. It is concluded from sequence and mutant studies that conserved amino acid residues around the histidyl-FAD are important for FAD binding; however, amino acids located more than 100 residues downstream from the histidyl in the Fp subunit can also effect<br/>flavinylation.},
  author       = {Maguire, John J. and Magnusson, Kerstin and Hederstedt, Lars},
  issn         = {0006-2960},
  language     = {eng},
  number       = {18},
  pages        = {5202--5208},
  publisher    = {The American Chemical Society},
  series       = {Biochemistry},
  title        = {<em>Bacillus subtilis</em> mutant succinate dehydrogenase lacking covalently bound flavin: Identification of the primary defect and studies on the iron-sulfur clusters in mutated and wild- type enzyme},
  url          = {http://dx.doi.org/10.1021/bi00366a033},
  volume       = {25},
  year         = {1986},
}