Initial characterization of site-directed mutants of tyrosine M210 in the reaction centre of Rhodobacter sphaeroides.
A description of the properties of site-directed mutants of the reaction centre (RC) of Rhodobacter sphaeroides is presented. The residue tyrosine M210 (YM210) has been changed to phenylalanine (FM210) and leucine (LM210). Both mutants grew photoheterotrophically under conditions of high light but only the FM210 mutant grew under low light. Photobleaching spectra of chromatophores isolated from these mutants showed that the amount of functional RC was comparable to wild-type and that the spectral features were essentially unchanged. Shifts were observed in the absorption spectra in regions attributable to all the chromophores. An increase in intensity and a 3 nm red shift (from 803 to 806 nm) was observed in the Qy band of the monomer bacteriochlorophylls. A new extinction coefficient for the RC was determined at 806 nm (332 +/- 15 mM-1 cm-1). Linear dichroism (LD) spectra showed that there was no significant large scale change in the angles of the individual pigments relative to the C2 axis of symmetry. Cytochrome turnover assays were performed on isolated RC and light harvesting I complex (LH I)-RC (photosynthetic units, PSU) preparations. A turnover number of 120 cyt RC-1 s-1 was calculated for both the mutants while wild-type had a turnover number of 228 cyt RC-1 s-1. The cytochrome c2-mediated re-reduction kinetics of P+ were comparable to those observed in the wild-type. The half-time of charge recombination within the RC increased in the mutants to the wild-type (100 ms in the wild type, and 150 and 200 ms in FM210 and LM210, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)
Gray
K A
KA
Department of Membrane Biochemistry, Max-Planck-Institut für Biochemie, Martinsried bei München, FRG.
Farchaus
J W
JW
Wachtveitl
J
J
Breton
J
J
Oesterhelt
D
D
D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D003574:Cytochrome c Group / Q000378:metabolism; D007930:Leucine; D008969:Molecular Sequence Data; D009154:Mutation; D015345:Oligonucleotide Probes; D010084:Oxidation-Reduction; D010649:Phenylalanine; D010788:Photosynthesis; D045322:Photosynthetic Reaction Center Complex Proteins; D015183:Restriction Mapping; D012242:Rhodobacter sphaeroides / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D013053:Spectrophotometry; D014443:Tyrosine
D001426:Bacterial Proteins; D003574:Cytochrome c Group; D015345:Oligonucleotide Probes; D045322:Photosynthetic Reaction Center Complex Proteins; D014443:Tyrosine; D010649:Phenylalanine; D007930:Leucine