THE A(T) THERMOLUMINESCENCE BAND FROM CHLAMYDOMONAS-REINHARDTII AND THE EFFECTS OF MUTAGENESIS OF HISTIDINE-RESIDUES ON THE DONOR SIDE OF THE PHOTOSYSTEM-II D1 POLYPEPTIDE

A(T) thermoluminescence bands were measured from thylakoids and Photosystem II (PS II) particles of wild-type (WT) and site-directed mutants of residues 195 (H195) and 190 (H190) of the PS II D1 polypeptide of Chlamydomonas reinhardtii. In wild type thylakoids the peak temperature of the A(T) band (pH 6.5) was at - 16.2 +/- 0.1 degrees C. Maximal thermoluminescence yield was achieved at an illumination temperature of approx. -20 degrees C, as previously observed in spinach PS II particles by Ono and Inoue (FEBS Lett. 278 (1991) 183-186). In contrast to previous observations, we found only a weak pH-dependence of the A, band intensity, from pH 5.5 to 8 in thylakoids or PS II particles from C. reinhardtii or spinach. Conversion of H195 of the D1 polypeptide of PS II to asparagine (H195N), tyrosine (H195Y), or aspartate (H195D) decreased the amplitudes of the A(T) bands to approx. 65, 53 and 38% of that of wild type, but did not significantly alter the peak temperatures of the bands. The residual A(T) bands in the H195 mutant strains showed illumination temperature- and flash number-dependencies, and pH dependencies similar to those of wild type. Therefore, we suggest that the quantum yield of the luminescence, rather than the energetics of the recombinatory pathway, was modified in the H195 mutants. In other experiments (Roffey, R.A., Kramer, D.M., Govindjee, and Sayre, R.T. (1994) Biochim. Biophys. Acta, in press), we concluded that the equilibrium constant between YZP+ and (YP)-P-Z+ in Mn-depleted material was dramatically shifted (about 50-fold) in the H195D mutant relative to WT. Since we found no effect of the H195 mutation on the peak temperature of the A(T) band, we concluded that Y-Z is probably not the trap for the positive charge involved in the A(T) band recombination, in agreement with the conclusion of Ono and Inoue (FEBS Lett. 278 (1991) 183-186). We also conclude that H195 cannot be the redox group responsible for carrying the positive charge for the AT band recombination. Modification of H190 to phenylalanine completely abolished the A(T) band, but also severely affected PS II donor reactions by slowing the electron transfer from tyrosine Y-Z to the oxidized primary donor P-680(+) by approx. 100- to 1000-fold. It was not possible to determine whether the loss of the A(T) band upon H190 modification was due to the removal of a redox-active histidine or to the secondary effects the modification had on electron transfer reactions. We present a kinetic model that explains many of the data on the A(T) band.