Mv. Ponamarev et Wa. Cramer, Perturbation of the internal water chain in cytochrome f of oxygenic photosynthesis: Loss of the concerted reduction of cytochromes f and b(6), BIOCHEM, 37(49), 1998, pp. 17199-17208
The 1.96 Angstrom structure of turnip cytochrome f revealed a linear intern
al chain of H2O molecules with the oxygen atoms of the chain having occupan
cies and "B" factors comparable to those of neighboring atoms [Martinez et
al, (1996) Protein Sci. 5, 1081-1092.]. Four waters extend 11 Angstrom from
the heme toward Lys66 on the cytochrome surface. All residues that contrib
ute an atom to the 15 H-bonds of five internal H2O molecules are essentiall
y conserved in 23 cytochrome sequences. With only Gin and Asn side chains i
nvolved in H-bonding, the water chain resembles a "proton wire". The functi
on of the conserved H2O chain was tested through site-directed mutagenesis
of these Asn and Gin residues. Four of the five conserved Asn/Gln residues
were changed in six mutants generated in the green alga, Chlamydomonas rein
hardtii. Except for the N168F mutant, all grew photosynthetically. Although
the rates of oxidation of cyt f oxidation and of reduction of cyt b6 (5-6
ms in the wild type) were not significantly affected, the rates of cyt f re
duction and generation of the slow electrochromic band shift (Delta psi(s))
were markedly decreased, the half-times increasing to as much as 38 and 18
ms, respectively. Thus, in these mutants, reduction of cyt bg reduction cl
early precedes that of cyt f, Retardation of Delta psi(s) in the absence of
an observable change in the rate of cyt bg reduction implied that the rate
of Hi translocation decreased in the mutants, and electron transfer was co
ncomitantly retarded, most likely between the ISP and cyt f. The following
was concluded: (i) proton and electron transfer are coupled in reduction of
cyt f, and the cyt f water chain functions in H+ transfer; (ii) reduction
of the high- and low-potential chains in the b(6)f complex is not concerted
in the water chain mutants; and (iii) quinol deprotonation and electron tr
ansfer from reduced quinone are initiated by an early event, probably the m
ovement of the ISP triggered by oxidation of cyt f.