H135A controls the redox activity of the Sco copper center. Kinetic and spectroscopic studies of the His135Ala variant of Bacillus subtilis Sco.
Sco-like proteins contain copper bound by two cysteines and a histidine residue. Although their function is still incompletely understood, there is a clear involvement with the assembly of cytochrome oxidases that contain the Cu(A) center in subunit 2, possibly mediating the transfer of copper into the Cu(A) binuclear site. We are investigating the reaction chemistry of BSco, the homologue from Bacillus subtilis. Our studies have revealed that BSco behaves more like a redox protein than a metallochaperone. The essential H135 residue that coordinates copper plays a role in stabilizing the Cu(II) rather than the Cu(I) form. When H135 is mutated to alanine, the oxidation rate of both hydrogen peroxide and one-electron outer-sphere reductants increases by 3 orders of magnitude, suggestive of a redox switch mechanism between the His-on and His-off conformational states of the protein. Imidazole binds to the H135A protein, restoring the N superhyperfine coupling in the EPR, but is unable to rescue the redox properties of wild-type Sco. These findings reveal a unique role for H135 in Sco function. We propose a hypothesis that electron transfer from Sco to the maturing oxidase may be essential for proper maturation and/or protection from oxidative damage during the assembly process. The findings also suggest that interaction of Sco with its protein partner(s) may perturb the Cu(II)-H135 interaction and thus induce a sensitive redox activity to the protein.
Siluvai
Gnana S
GS
Department of Science and Engineering, Oregon Health and Science University, Beaverton, Oregon 97006, USA.
Nakano
Michiko M
MM
Mayfield
Mary
M
Nilges
Mark J
MJ
Blackburn
Ninian J
NJ
D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D003300:Copper / Q000737:chemistry* / Q000378:metabolism; D014644:Genetic Variation; D007700:Kinetics; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D020125:Mutation, Missense; D010084:Oxidation-Reduction; D013057:Spectrum Analysis
D001426:Bacterial Proteins; D008565:Membrane Proteins; C482909:Sco protein, Bacillus subtilis; D003300:Copper