TY - JOUR
T1 - Spectroscopic and DFT studies of second-sphere variants of the type 1 copper site in azurin
T2 - Covalent and nonlocal electrostatic contributions to reduction potentials
AU - Hadt, Ryan G.
AU - Sun, Ning
AU - Marshall, Nicholas M.
AU - Hodgson, Keith O.
AU - Hedman, Britt
AU - Lu, Yi
AU - Solomon, Edward I.
PY - 2012/10/10
Y1 - 2012/10/10
N2 - The reduction potentials (E0) of type 1 (T1) or blue copper (BC) sites in proteins and enzymes with identical first coordination spheres around the redox active copper ion can vary by ∼400 mV. Here, we use a combination of low-temperature electronic absorption and magnetic circular dichroism, electron paramagnetic resonance, resonance Raman, and S K-edge X-ray absorption spectroscopies to investigate a series of second-sphere variants-F114P, N47S, and F114N in Pseudomonas aeruginosa azurin-which modulate hydrogen bonding to and protein-derived dipoles nearby the Cu-S(Cys) bond. Density functional theory calculations correlated to the experimental data allow for the fractionation of the contributions to tuning E0 into covalent and nonlocal electrostatic components. These are found to be significant, comparable in magnitude, and additive for active H-bonds, while passive H-bonds are mostly nonlocal electrostatic in nature. For dipoles, these terms can be additive to or oppose one another. This study provides a methodology for uncoupling covalency from nonlocal electrostatics, which, when coupled to X-ray crystallographic data, distinguishes specific local interactions from more long-range protein/active interactions, while affording further insight into the second-sphere mechanisms available to the protein to tune the E0 of electron-transfer sites in biology.
AB - The reduction potentials (E0) of type 1 (T1) or blue copper (BC) sites in proteins and enzymes with identical first coordination spheres around the redox active copper ion can vary by ∼400 mV. Here, we use a combination of low-temperature electronic absorption and magnetic circular dichroism, electron paramagnetic resonance, resonance Raman, and S K-edge X-ray absorption spectroscopies to investigate a series of second-sphere variants-F114P, N47S, and F114N in Pseudomonas aeruginosa azurin-which modulate hydrogen bonding to and protein-derived dipoles nearby the Cu-S(Cys) bond. Density functional theory calculations correlated to the experimental data allow for the fractionation of the contributions to tuning E0 into covalent and nonlocal electrostatic components. These are found to be significant, comparable in magnitude, and additive for active H-bonds, while passive H-bonds are mostly nonlocal electrostatic in nature. For dipoles, these terms can be additive to or oppose one another. This study provides a methodology for uncoupling covalency from nonlocal electrostatics, which, when coupled to X-ray crystallographic data, distinguishes specific local interactions from more long-range protein/active interactions, while affording further insight into the second-sphere mechanisms available to the protein to tune the E0 of electron-transfer sites in biology.
UR - http://www.scopus.com/inward/record.url?scp=84961982126&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84961982126&partnerID=8YFLogxK
U2 - 10.1021/ja306438n
DO - 10.1021/ja306438n
M3 - Article
C2 - 22985400
AN - SCOPUS:84961982126
SN - 0002-7863
VL - 134
SP - 16701
EP - 16716
JO - Journal of the American Chemical Society
JF - Journal of the American Chemical Society
IS - 40
ER -