Effects of the Tyr64 Substitution on the Stability of Cytochrome c₅₅₃, a low Oxidoreduction‐Potential Cytochrome from Desulfovibrio vulgaris Hildenborough

Abstract

Cytochrome c553 from sulfate‐reducing bacteria is a lowoxidoreduction‐potential cytochrome. The primary and tertiary structures show notable differences when compared to mitochondrial cytochromes. Tyr64 replacement in cytochrome c553 provides evidence that this residue is not directly involved in the potential modulation but is mostly implicated in the hydrogenbond network around the heme. While the different variants obtained did not induce drastic structural modifications, they did affect the stability of the protein. This decrease of stability in acidic and alkaline environments was observed by variations in the optical spectra and by mass spectrometry. In addition, the mobility of aromatic side‐chain was found to be increased in the mutant proteins as monitored by two‐dimensional NMR spectroscopy.

New conformational properties induced by the replacement of Tyr‐64 in Desulfovibrio vulgaris Hildenborough ferricytochrome c553 using isotopic exchanges monitored by mass spectrometry

“…DvH wild-type and muAbbreviations." ESI, electrospray ionization; MALDI, matrix-assisted tants ferricytochrome e~,~:~ were obtained as previously described laser desorption/ionization; H/D, hydrogen/deuterium; DvH Desu(fb- [24,25]. vibrio vulgaris Hildenborough; LC, liquid chromatography; MS, mass spectrometry.…”

Stability study of Rhodobacter capsulatus ferrocytochrome c2 wild‐type and site‐directed mutants using hydrogen/deuterium exchange monitored by electrospray ionization mass spectrometry

“…Site-directed mutants have been generated in a number of species including: Rhodobacter capsulatus cytochrome c2 [17], Thiobacillus versutus cytochrome C55o [18], DesulJovibrio vulgaris cytochrome c553 [19], Saccharomyces cerevisiae (yeast) cytochrome c [20], Drosophila melanogaster (fruit fly) cytochrome c [21], and Rattus norvegicus (rat) cytochrome c [22]. In the present study, the stability properties of Rb.…”

15N‐labelling and preliminary heteronuclear NMR study ofDesulfovibrio vulgaris Hildenborough cytochrome c553

“…We have already described that this minor conformation remains during the reduction of the cytochrome, and the ratio of the two conformations decreases with time in the reduced state [34]. MS has provided evidence that three conformers of cytochrome c 553 exist in a dynamic equilibrium with a mass increase of +16 and +32 Da [35]. This increase in mass has been correlated to an oxidative damage to the methionine residue in a sulphoxide or sulphone derivative.…”