Gene Expression in Rhodopseudomonas Sphaeroides (Photosynthesis, Carboxylase, Membrane Biosynthesis
A DNA-directed coupled transcription-translation system has been developed in cell-free extracts from the facultative phototroph, Rhodopseudomonas sphaeroides. The in vitro protein synthesizing system was active when prepared from either aerobically or anaerobically grown cells. Optimal activity was dependent upon use of extracts prepared freshly from early exponential phase cells, the method of cell breakage, and the length of preincubation of the extract (S-30), as well as the concentrations of S-30, DNA template, and cations. The cell-free system was used to study the physical and biochemical characterization of an enzymatically active form II ribulose-1,5-bisphosphate carboxylase which was synthesized and assembled in vitro from cloned R. sphaeroides DNA. The predominant enzymatically active oligomeric form of the in vitro synthesized and assembled RuBP carboxylase was a tetramer-hexamer, and assembly of the monomeric subunits to form active enzyme was dependent on the presence of a minimum number of subunits.
University of Illinois - Urbana-Champaign
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University of Illinois at Urbana-Champaign