Site-directed mutations at D1-His198 and D2-His 97 of photosystem II in synechocystis PCC 6803: Sites of primary charge separation and cation and triplet stabilization
Ba. Diner et al., Site-directed mutations at D1-His198 and D2-His 97 of photosystem II in synechocystis PCC 6803: Sites of primary charge separation and cation and triplet stabilization, BIOCHEM, 40(31), 2001, pp. 9265-9281
Site-directed mutations were introduced to replace D1-His198 and D2-His197
of the Dl and D2 polypeptides, respectively, of the photosystem II (PSII) r
eaction center of Synechocystis PCC 6803. These residues coordinate chlorop
hylls P-A and P-B which are homologous to the special pair Bchlorophylls of
the bacterial reaction centers that are coordinated respectively by histid
ines L-173 and M-200 (202). P-A and P-B together serve as the primary elect
ron donor, P, in purple bacterial reaction centers. In PS II, the site-dire
cted mutations at D1 His198 affect the P+-P-absorbance difference spectrum.
The bleaching maximum in the Soret region (in WT at 433 nm) is blue-shifte
d by as much as 3 nm. In the D1 His 198Gln mutant, a similar displacement t
o the blue is observed for the bleaching maximum in the Q(y) region (672.5
nm in WT at 80 K), whereas features attributed to a band shift centered at
681 nm are not altered. In the Y-z. -Y-z-difference spectrum, the band shif
t of a reaction center chlorophyll centered in WT at 433-434 nm is shifted
by 2-3 nm to the blue in the D1-His198Gln mutant. The D1-His198Gln mutation
has little effect on the optical difference spectrum, P-3-P-1, of the reac
tion center triplet formed by P(+)Pheo(-) charge recombination (bleaching a
t 681-684 nm), measured at 5-80 K, but becomes visible as a pronounced shou
lder at 669 nm at temperatures greater than or equal to 150 K. Measurements
of the kinetics of oxidized donor-Q(A)(-) charge recombination and of the
reduction of P+ by redox active tyrosine, Y-z, indicate that the reduction
potential of the redox couple P+/P can be appreciably modulated both positi
vely and negatively by ligand replacement at D1-198 but somewhat less so at
D2-197. On the basis of these observations and others in the literature, w
e propose that the monomeric accessory chlorophyll, B-A, is a long-waveleng
th trap located at 684 nm at 5 K. B-A* initiates primary charge separation
at low temperature, a function that is increasingly shared with P-A* in an
activated process as the temperature rises. Charge separation from B-A* wou
ld be potentially very fast and form PA+BA- and/or B(A)(+)Pheo(-) as observ
ed in bacterial reaction centers upon direct excitation of B-A (van Bredero
de, M. E., et al. (1999) Proc. Nad. Acad Sci. 96, 2054-2059). The cation, g
enerated upon primary charge separation in PSII, is stabilized at all tempe
ratures primarily on P-A, the absorbance spectrum of which is displaced to
the blue by the mutations. In WT, the cation is proposed to be shared to a
minor extent (similar to 20%) with P-B, the contribution of which can be mo
dulated up or down by mutation. The band shift at 681 nm, observed in the P
+-P difference spectrum, is attributed to an electrochromic effect of P-A() on neighboring B-A. Because of its low-energy singlet and therefore tripl
et state, the reaction center triplet state is stabilized on B-A at less th
an or equal to 80 K but can be shared with P-A at > 80 K in a thermally act
ivated process.