Direct and mediated electron transfer between intact succinate:quinone oxidoreductase from Bacillus subtilis and a surface modified gold electrode reveals redox state-dependent conformational changes

TY - JOUR

T1 - Direct and mediated electron transfer between intact succinate:quinone oxidoreductase from Bacillus subtilis and a surface modified gold electrode reveals redox state-dependent conformational changes

AU - Christenson, Andreas

AU - Gustavsson, Tobias

AU - Gorton, Lo

AU - Hägerhäll, Cecilia

PY - 2008/9

Y1 - 2008/9

N2 - Succinate:quinone oxidoreductase (SQR) from Bacillus subtilis consists of two hydrophilic protein subunits comprising succinate dehydrogenase, and a di-heme membrane anchor protein harboring two putative quinone binding sites, Q(p) and Q(d). In this work we have used spectroelectrochemistry to study the electronic communication between purified SQR and a surface modified gold capillary electrode. In the presence of two soluble quinone mediators the midpoint potentials of both hemes were revealed essentially as previously determined by conventional redox titration (heme b(H), E(m)=+65 mV, heme b(L), E(m)=-95 mV). In the absence of mediators the enzyme still communicated with the electrode, albeit with a reproducible hysteresis, resulting in the reduction of both hemes occurring approximately at the midpoint potential of heme b(L), and with a pronounced delay of reoxidation. When the specific inhibitor 2-n-heptyl-4 hydroxyquinoline N-oxide (HQNO), which binds to Q(d) in B. subtilis SQR, was added together with the two quinone mediators, rapid reductive titration was still possible which can be envisioned as an electron transfer occurring via the HQNO insensitive Q(p) site. In contrast, the subsequent oxidative titration was severely hampered in the presence of HQNO, in fact it completely resembled the unmediated reaction. If mediators communicate with Q(p) or Q(d), either event is followed by very rapid electron redistribution within the enzyme. Taken together, this strongly suggests that the accessibility of Q(p) depended on the redox state of the hemes. When both hemes were reduced, and Q(d) was blocked by HQNO, quinone-mediated communication via the Q(p) site was no longer possible, revealing a redox-dependent conformational change in the membrane anchor domain.

AB - Succinate:quinone oxidoreductase (SQR) from Bacillus subtilis consists of two hydrophilic protein subunits comprising succinate dehydrogenase, and a di-heme membrane anchor protein harboring two putative quinone binding sites, Q(p) and Q(d). In this work we have used spectroelectrochemistry to study the electronic communication between purified SQR and a surface modified gold capillary electrode. In the presence of two soluble quinone mediators the midpoint potentials of both hemes were revealed essentially as previously determined by conventional redox titration (heme b(H), E(m)=+65 mV, heme b(L), E(m)=-95 mV). In the absence of mediators the enzyme still communicated with the electrode, albeit with a reproducible hysteresis, resulting in the reduction of both hemes occurring approximately at the midpoint potential of heme b(L), and with a pronounced delay of reoxidation. When the specific inhibitor 2-n-heptyl-4 hydroxyquinoline N-oxide (HQNO), which binds to Q(d) in B. subtilis SQR, was added together with the two quinone mediators, rapid reductive titration was still possible which can be envisioned as an electron transfer occurring via the HQNO insensitive Q(p) site. In contrast, the subsequent oxidative titration was severely hampered in the presence of HQNO, in fact it completely resembled the unmediated reaction. If mediators communicate with Q(p) or Q(d), either event is followed by very rapid electron redistribution within the enzyme. Taken together, this strongly suggests that the accessibility of Q(p) depended on the redox state of the hemes. When both hemes were reduced, and Q(d) was blocked by HQNO, quinone-mediated communication via the Q(p) site was no longer possible, revealing a redox-dependent conformational change in the membrane anchor domain.

KW - Bacillus subtilis/drug effects

KW - Catalysis/drug effects

KW - Electrochemistry

KW - Electrodes

KW - Electron Transport/drug effects

KW - Electron Transport Complex II/chemistry

KW - Gold/metabolism

KW - Heme/chemistry

KW - Hydroxyquinolines/pharmacology

KW - Models, Biological

KW - Oxidation-Reduction/drug effects

KW - Protein Structure, Secondary

KW - Spectrum Analysis

U2 - 10.1016/j.bbabio.2008.05.450

DO - 10.1016/j.bbabio.2008.05.450

M3 - Journal article

C2 - 18598669

SN - 0006-3002

VL - 1777

SP - 1203

EP - 1210

JO - B B A - Reviews on Cancer

JF - B B A - Reviews on Cancer

IS - 9

ER -