ABSTRACT
Bacillus megaterium accumulates poly-(3-hydroxybutyrate) (PHB) as a reserve material in intracellular granules. In this work we described a method for the preparation of PHB granules from B. megaterium PV447 and the analysis by polyacrylamide gel electrophoresis of the associated proteins. By comparison with another species a function is proposed for some of these proteins.
Subject(s)
Bacillus megaterium/chemistry , Bacterial Proteins/isolation & purification , Cytoplasmic Granules/chemistry , Hydroxybutyrates/analysis , Plant Lectins , Polyesters/analysis , Acyltransferases/isolation & purification , Bacillus megaterium/enzymology , Bacillus megaterium/ultrastructure , Cell Fractionation , Chromatium/enzymology , Electrophoresis, Polyacrylamide Gel , Lectins/isolation & purification , Species SpecificityABSTRACT
We have recently reported the expression in Escherichia coli of an enzymatically competent ferredoxin-NADP+ oxidoreductase from cloned pea genes encoding either the mature enzyme or its precursor protein (Ceccarelli, E. A., Viale, A. M., Krapp, A. R., and Carrillo, N. (1991) J. Biol. Chem. 266, 14283-14287). Processing to the mature form by bacterial protease(s) and FAD assembly occurred in the bacterial cytosol. Expression of ferredoxin-NADP+ reductase in chaperonin-deficient (groE-) mutants of E. coli resulted in partial reductase assembly at permissive growth temperatures (i.e. 30 degrees C), and in total breakdown of holoenzyme assembly, and accumulation as aggregated inclusion bodies at non-permissive temperatures (i.e. 42 degrees C). Coexpression in these mutants of a cloned groESL operon from the phototrophic bacterium Chromatium vinosum resulted in partial or total recoveries of ferredoxin-NADP+ reductase assembly. The overall results indicate that bacterial chaperonins are required for the productive folding/assembly of eucaryotic ferredoxin-NADP+ reductase expressed in E. coli.
Subject(s)
Bacterial Proteins/metabolism , Chaperonins/metabolism , Chromatium/genetics , Escherichia coli/genetics , Ferredoxin-NADP Reductase/genetics , Operon , Bacterial Proteins/genetics , Chaperonin 10 , Chaperonin 60 , Chromatium/enzymology , Cloning, Molecular , Escherichia coli/metabolism , Escherichia coli Proteins , Gene Expression , Heat-Shock Proteins/genetics , PlasmidsABSTRACT
An open reading frame, rbcR, was identified 226 bp upstream of rbcAB, i.e., the ribulose 1,5-bisphosphate carboxylase genes expressed in the phototrophic purple bacterium Chromatium vinosum. Several features reveal that rbcR encodes a member of the LysR family of transcriptional regulators, in which an anomalous content of lysine and arginine residues (Lys/Arg anomaly) was found. The expression of rbcR in Escherichia coli as a protein fused to the N-terminal region of beta-galactosidase led to reduced expression of rbcAB. Thus, rbcR is likely to encode a trans-acting transcriptional regulator of rbcAB expression in C. vinosum.