Echalier, Aude, Brittain, Thomas, Wright, Joshua, Boycheva, Svetlana, Mortuza, Gulnahar B., Fulop, Vilmos and Watmough, Nicholas J.. (2008) Redox-linked structural changes associated with the formation of a catalytically competent form of the diheme cytochrome c peroxidase from Pseudomonas aeruginosa. Biochemistry, Vol.47 (No.7). pp. 1947-1956. ISSN 0006-2960

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Abstract

A recombinant form of the prototypic diheme bacterial cytochrome c peroxidase (BCCP) from Pseudomonas aeruginosa (PsaCCP) has been expressed in Escherichia coli and purified to homogeneity. This material was used to carry out the first integrated biochemical, spectroscopic and structural investigation of the factors leading to reductive activation of this class of enzymes. A single, tightly bound, Ca2+ ion (K = 3 x 10(10) M-1) found at the domain interface of both the fully oxidized and mixed-valence forms of the enzyme is absolutely required for catalytic activity. Reduction of the electron-transferring (high-potential) heme in the presence of Ca2+ ions triggers substantial structural rearrangements around the active-site (low-potential) heme to allow substrate binding and catalysis. The enzyme also forms a mixed-valence state in the absence of Ca2+ ions, but a combination of electronic absorption, and EPR spectroscopies suggests that under these circumstances the low potential heme remains six-coordinate, unable to bind substrate and therefore catalytically inactive. Our observations strongly suggest that the two mixed-valence forms of native PsaCCP reported previously by Foote and colleagues (Foote, N., Peterson, J., Gadsby, P., Greenwood, C., and Thomson, A. (1985) Biochem. J. 230, 227-237) correspond to the Ca2+-loaded and -depleted forms of the enzyme.

Item Type:	Journal Article
Subjects:	Q Science > QD Chemistry
Divisions:	Faculty of Science > Life Sciences (2010- ) > Biological Sciences ( -2010)
Journal or Publication Title:	Biochemistry
Publisher:	American Chemical Society
ISSN:	0006-2960
Date:	19 February 2008
Volume:	Vol.47
Number:	No.7
Number of Pages:	10
Page Range:	pp. 1947-1956
Identification Number:	10.1021/bi702064f
Status:	Peer Reviewed
Publication Status:	Published
Access rights to Published version:	Restricted or Subscription Access
URI:	http://wrap.warwick.ac.uk/id/eprint/30589

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