TY - JOUR
T1 - The selenocysteine-substituted blue copper center
T2 - Spectroscopic investigations of Cys112SeCys Pseudomonas aeruginosa azurin
AU - Ralle, Martina
AU - Berry, Steven M.
AU - Nilges, Mark J.
AU - Gieselman, Matt D.
AU - Van Der Donk, Wilfred A.
AU - Lu, Yi
AU - Blackburn, Ninian J.
PY - 2004/6/16
Y1 - 2004/6/16
N2 - Azurin is a small electron-transfer protein belonging to the cupredoxin family. The Cu atom is located within a trigonal plane coordinated by two histidines (His46 and His117) and a cysteine (Cys112) with two more distant ligands (Gly45 and Met121) providing axial interactions. A Cys112SeCys derivative has been prepared by expressed protein ligation, and detailed UV/vis, EPR and EXAFS studies at the Cu and Se K-edges have been carried out. Marked changes are observed between the EPR parameters of the Cys112SeCys and WT azurin derivatives, which include a 2-fold increase in A11, a decrease in g-values, and a large increase in rhombicity of the g-tensor. The Cu-Se and Se-Cu bond lengths obtained from analysis of the Cu and Se K-EXAFS of the oxidized protein were found to be 2.30 and 2.31 Å, respectively, 0.14 Å longer than the Cu-S distance of the WT protein. Unexpectedly, the Cu-Se bond lengths were found to undergo only minor changes during reduction, suggesting a very similar structure in both redox states and extending the "rack" hypothesis to the Se-substituted protein.
AB - Azurin is a small electron-transfer protein belonging to the cupredoxin family. The Cu atom is located within a trigonal plane coordinated by two histidines (His46 and His117) and a cysteine (Cys112) with two more distant ligands (Gly45 and Met121) providing axial interactions. A Cys112SeCys derivative has been prepared by expressed protein ligation, and detailed UV/vis, EPR and EXAFS studies at the Cu and Se K-edges have been carried out. Marked changes are observed between the EPR parameters of the Cys112SeCys and WT azurin derivatives, which include a 2-fold increase in A11, a decrease in g-values, and a large increase in rhombicity of the g-tensor. The Cu-Se and Se-Cu bond lengths obtained from analysis of the Cu and Se K-EXAFS of the oxidized protein were found to be 2.30 and 2.31 Å, respectively, 0.14 Å longer than the Cu-S distance of the WT protein. Unexpectedly, the Cu-Se bond lengths were found to undergo only minor changes during reduction, suggesting a very similar structure in both redox states and extending the "rack" hypothesis to the Se-substituted protein.
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U2 - 10.1021/ja031821h
DO - 10.1021/ja031821h
M3 - Article
C2 - 15186162
AN - SCOPUS:2942595805
SN - 0002-7863
VL - 126
SP - 7244
EP - 7256
JO - Journal of the American Chemical Society
JF - Journal of the American Chemical Society
IS - 23
ER -