Design of a ruthenium-labeled cytochrome c derivative to study electron transfer with the cytochrome bc1 complex.
A new ruthenium-cytochrome c derivative was designed to study electron transfer from cytochrome bc1 to cytochrome c (Cc). The single sulfhydryl on yeast H39C;C102T iso-1-Cc was labeled with Ru(2,2'-bipyrazine)2(4-bromomethyl-4'-methyl-2,2'-bipyridine) to form Ru(z)-39-Cc. The Ru(z)-39-Cc derivative has the same steady-state activity with yeast cytochrome bc1 as wild-type yeast iso-1-Cc, indicating that the ruthenium complex does not interfere in the binding interaction. Laser excitation of reduced Ru(z)-39-Cc results in electron transfer from heme c to the excited state of ruthenium with a rate constant of 1.5 x 10(6) x s(-1). The resulting Ru(I) is rapidly oxidized by atmospheric oxygen in the buffer. The yield of photooxidized heme c is 20% in a single flash. Flash photolysis of a 1:1 complex between reduced yeast cytochrome bc1 and Ru(z)-39-Cc at low ionic strength leads to rapid photooxidation of heme c, followed by intracomplex electron transfer from cytochrome c1 to heme c with a rate constant of 1.4 x 10(4) x s(-1). As the ionic strength is raised above 100 mM, the intracomplex phase disappears, and a new phase appears due to the bimolecular reaction between solution Ru-39-Cc and cytochrome bc1. The interaction of yeast Ru-39-Cc with yeast cytochrome bc1 is stronger than that of horse Ru-39-Cc with bovine cytochrome bc1, suggesting that nonpolar interactions are stronger in the yeast system.
Engstrom
Gregory
G
Department of Chemistry and Biochemistry, University of Arkansas, Fayetteville, Arkansas 72701, USA.
Rajagukguk
Ray
R
Saunders
Aleister J
AJ
Patel
Chetan N
CN
Rajagukguk
Sany
S
Merbitz-Zahradnik
Torsten
T
Xiao
Kunhong
K
Pielak
Gary J
GJ
Trumpower
Bernard
B
Yu
Chang-An
CA
Yu
Linda
L
Durham
Bill
B
Millett
Francis
F
D018360:Crystallography, X-Ray; D003574:Cytochrome c Group / Q000138:chemical synthesis*; D004579:Electron Transport; D014450:Electron Transport Complex III / Q000737:chemistry*; D006418:Heme / Q000031:analogs & derivatives* / Q000737:chemistry; D007700:Kinetics; D008956:Models, Chemical; D009942:Organometallic Compounds / Q000138:chemical synthesis; D009994:Osmolar Concentration; D010782:Photolysis; D012428:Ruthenium / Q000737:chemistry*; D029701:Saccharomyces cerevisiae Proteins / Q000138:chemical synthesis* / Q000737:chemistry
C091938:(4-bromomethyl-4'-methylbipyridine)(bis(bipyridine))ruthenium(II); D003574:Cytochrome c Group; D009942:Organometallic Compounds; D029701:Saccharomyces cerevisiae Proteins; C031690:heme C; D006418:Heme; D012428:Ruthenium; D014450:Electron Transport Complex III