Thermal Instability of Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase from a Temperature-Conditional Chloroplast Mutant of Chlamydomonas reinhardtii.
Mutant 68-4PP of Chlamydomonas reinhardtii has only 10% of the normal level of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) holoenzyme when grown at 35[deg]C. However, when grown at 25[deg]C, the amount of holoenzyme is greater than 35% of the wild-type level, and the purified enzyme has a reduced CO2/O2 specificity factor. These mutant characteristics result from a chloroplast mutation that causes leucine-290 to be replaced by phenylalanine within the Rubisco large-subunit protein. A nuclear mutation (named S52-2B) was previously identified that can suppress both the in vivo instability and reduced CO2/O2 specificity of the mutant enzyme. However, the effect of this nuclear mutation on the in vitro stability of the holoenzyme was not resolved. In the present study, purified Rubisco from mutant 68-4PP was found to be less thermally stable than the wild-type enzyme, and it had maximal carboxylase activity at a lower temperature. When incubated at 35[deg]C, the mutant enzyme lost carboxylase activity at a much faster rate than the wild-type enzyme. However, the nuclear S52-2B suppresor mutation improved the thermal stability of the mutant enzyme in all cases. These results indicate that structural changes in mutant 68-4PP Rubisco can account for its observed inactivation in vitro and degradation in vivo. Such structural alterations are alleviated by the function of a nuclear gene.
Chen
Z.
Z
Department of Biochemistry, University of Nebraska, Lincoln, Nebraska 68583-0718.
Hong
S.
S
Spreitzer
R. J.
RJ