Mutations designed to modify the environment of the primary electron donor of the reaction center from Rhodobacter sphaeroides: phenylalanine to leucine at L167 and histidine to phenylalanine at L168.
Two mutations, L168 His to Phe and L167 Phe to Leu, were made in residues near the primary electron donor, a bacteriochlorophyll dimer, of the reaction center from Rhodobacter sphaeroides. Blue shifts of 10-15 nm in the 865-nm band of the donor were observed in the optical absorption spectra of both of the mutant reaction centers. The rate of initial electron transfer was determined by measurement of the kinetics of the decay of the excited state of the donor, and the rate of charge recombination was determined by measurement of the recovery of the bleaching of the donor. The initial electron transfer time constant and the charge recombination time constant were determined to be 3.6 ps and 220 ms, respectively, in the L168 His to Phe mutant and 5.0 ps and 85 ms in the L167 Phe to Leu mutant, compared to 3.8 ps and 100 ms measured for the wild type. The oxidation potential of the donor measured by oxidation-reduction titrations was found to decrease by 80 mV in the L168 His to Phe mutant and increase by 25 mV in the L167 Phe to Leu mutant. Time-resolved fluorescence decay measurements indicated that the change in the oxidation potential of the donor in the L168 His to Phe mutant resulted in a change in the energies of the charge-separated states. The results show that an increase in the driving force does not increase the rate of the initial electron transfer reaction.(ABSTRACT TRUNCATED AT 250 WORDS)
Murchison
H A
HA
Department of Chemistry and Biochemistry, Arizona State University, Tempe 85287-1604.
Alden
R G
RG
Allen
J P
JP
Peloquin
J M
JM
Taguchi
A K
AK
Woodbury
N W
NW
Williams
J C
JC
D006639:Histidine / Q000737:chemistry; D007930:Leucine / Q000737:chemistry; D045342:Light-Harvesting Protein Complexes; D016297:Mutagenesis, Site-Directed; D010084:Oxidation-Reduction; D010649:Phenylalanine / Q000737:chemistry; D010788:Photosynthesis; D045322:Photosynthetic Reaction Center Complex Proteins / Q000737:chemistry*; D012242:Rhodobacter sphaeroides / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013057:Spectrum Analysis; D013329:Structure-Activity Relationship
D045342:Light-Harvesting Protein Complexes; D045322:Photosynthetic Reaction Center Complex Proteins; D010649:Phenylalanine; D006639:Histidine; D007930:Leucine