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 Photos ystem 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 temperatur e of the A(T) band (pH 6.5) was at - 16.2 +/- 0.1 degrees C. Maximal t hermoluminescence yield was achieved at an illumination temperature of approx. -20 degrees C, as previously observed in spinach PS II partic les by Ono and Inoue (FEBS Lett. 278 (1991) 183-186). In contrast to p revious observations, we found only a weak pH-dependence of the A, ban d 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 P S II to asparagine (H195N), tyrosine (H195Y), or aspartate (H195D) dec reased the amplitudes of the A(T) bands to approx. 65, 53 and 38% of t hat of wild type, but did not significantly alter the peak temperature s of the bands. The residual A(T) bands in the H195 mutant strains sho wed illumination temperature- and flash number-dependencies, and pH de pendencies similar to those of wild type. Therefore, we suggest that t he quantum yield of the luminescence, rather than the energetics of th e recombinatory pathway, was modified in the H195 mutants. In other ex periments (Roffey, R.A., Kramer, D.M., Govindjee, and Sayre, R.T. (199 4) Biochim. Biophys. Acta, in press), we concluded that the equilibriu m constant between YZP+ and (YP)-P-Z+ in Mn-depleted material was dram atically shifted (about 50-fold) in the H195D mutant relative to WT. S ince we found no effect of the H195 mutation on the peak temperature o f the A(T) band, we concluded that Y-Z is probably not the trap for th e positive charge involved in the A(T) band recombination, in agreemen t 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. Modificat ion of H190 to phenylalanine completely abolished the A(T) band, but a lso severely affected PS II donor reactions by slowing the electron tr ansfer from tyrosine Y-Z to the oxidized primary donor P-680(+) by app rox. 100- to 1000-fold. It was not possible to determine whether the l oss 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 ex plains many of the data on the A(T) band.