Intermonomer electron transfer between the low-potential b hemes of cytochrome bc₁.
Cytochrome (cyt) bc(1) is a structural dimer with its monomers consisting of the Fe-S protein, cyt b, and cyt c(1) subunits. Its three-dimensional architecture depicts it as a symmetrical homodimer, but the mobility of the head domain of the Fe-S protein indicates that the functional enzyme exists in asymmetrical heterodimeric conformations. Here, we report a new genetic system for studying intra- and intermonomer interactions within the cyt bc(1) using the facultative phototrophic bacterium Rhodobacter capsulatus. The system involves two different sets of independently expressed cyt bc(1) structural genes carried by two plasmids that are coharbored by a cell without its endogenous enzyme. Our results indicate that coexpressed cyt bc(1) subunits were matured, assorted, and assembled in vivo into homo- and heterodimeric enzymes that can bear different mutations in each monomer. Using the system, the occurrence of intermonomer electron transfer between the low-potential b hemes of cyt bc(1) was probed by choosing mutations that perturb electron transfer at the hydroquinone oxidation (Q(o)) and quinone reduction (Q(i)) sites of the enzyme. The data demonstrate that active heterodimeric variants, formed of monomers carrying mutations that abolish only one of the two (Q(o) or Q(i)) active sites of each monomer, are produced, and they support photosynthetic growth of R. capsulatus. Detailed analyses of the physicochemical properties of membranes of these mutants, as well as purified homo- and heterodimeric cyt bc(1) preparations, demonstrated that efficient and productive electron transfer occurs between the low-potential b(L) hemes of the monomers in a heterodimeric enzyme. Overall findings are discussed with respect to intra- and intermonomer interactions that take place during the catalytic turnover of cyt bc(1).
Department of Biology, University of Pennsylvania, Philadelphia, Pennsylvania 19104, United States.
Pascal
P
Lanciano
Dong-Woo
DW
Lee
Honghui
H
Yang
Elisabeth
E
Darrouzet
Fevzi
F
Daldal
D004579:Electron Transport; D014450:Electron Transport Complex III / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006418:Heme / Q000737:chemistry* / Q000378:metabolism; D007700:Kinetics; D008958:Models, Molecular; D010084:Oxidation-Reduction; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D016354:Rhodobacter capsulatus / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
D006418:Heme; D014450:Electron Transport Complex III
Photosynth Res. 2004;81(3):251-75
16034531
Biochemistry. 1992 Apr 7;31(13):3342-51
1313292
Biochim Biophys Acta. 2002 Sep 10;1555(1-3):166-73
12206910
Biochemistry. 1994 Jan 25;33(3):723-33
8292600
Trends Biochem Sci. 2005 Apr;30(4):176-82
15817393
J Biol Chem. 1984 Feb 25;259(4):2331-6
6321467
FEBS J. 2007 Sep;274(17):4526-39
17680808
Anal Biochem. 1985 Oct;150(1):76-85
3843705
J Biol Chem. 2005 Mar 11;280(10):9251-7
15615714
Methods Enzymol. 1978;54:411-35
732578
Biochemistry. 1992 Apr 7;31(13):3351-8
1313293
FEBS Lett. 1998 Jul 17;431(2):291-6
9708922
Biochemistry. 2005 Aug 9;44(31):10520-32
16060661
Science. 1998 Jul 3;281(5373):64-71
9651245
Nature. 1970 Aug 15;227(5259):680-5
5432063
J Biol Chem. 2008 May 16;283(20):13973-82
18343816
Biochemistry. 2009 Mar 10;48(9):1888-99
19254042
Nature. 2004 Feb 12;427(6975):607-12
14961113
J Mol Biol. 1987 May 5;195(1):1-12
2821266
Proc Natl Acad Sci U S A. 2000 Apr 25;97(9):4567-72
10781061
Annu Rev Biochem. 2000;69:1005-75
10966481
Biochemistry. 2000 Dec 19;39(50):15475-83
11112533
Biochim Biophys Acta. 2008 Jul-Aug;1777(7-8):1001-19
18501698
J Biol Chem. 1990 Jul 15;265(20):11409-12
2164001
Biochemistry. 1998 Nov 17;37(46):16242-51
9819216
Nature. 1998 Apr 16;392(6677):677-84
9565029
J Biol Chem. 1986 Jan 15;261(2):584-91
3001072
J Biol Chem. 2010 Jan 1;285(1):502-10
19892700
Trends Biochem Sci. 2001 Jul;26(7):445-51
11440857
J Theor Biol. 1976 Oct 21;62(2):327-67
186667
Biochim Biophys Acta. 1983 May 23;723(2):202-218
21494412
Biochim Biophys Acta. 2010 Dec;1797(12):1842-8
20513347
J Biol Chem. 2008 Jun 20;283(25):17542-9
18390544
Science. 2010 Jul 23;329(5990):451-4
20651150
J Biol Chem. 2005 Jun 17;280(24):22732-40
15833742
Gene. 1977;2(2):95-113
344137
Proc Natl Acad Sci U S A. 1991 Jan 15;88(2):492-6
1846443
Biochemistry. 1995 Dec 12;34(49):16004-12
8519756
Biochemistry. 1999 Nov 30;38(48):15807-26
10625446
Science. 1997 Jul 4;277(5322):60-6
9204897
Biochemistry. 1998 Jun 2;37(22):8105-14
9609705