Chloroplast molecular chaperone-assisted refolding and reconstitution of an active multisubunit coupling factor CF1 core.
AUTOR(ES)
Chen, G G
RESUMO
The chloroplast coupling factor 1 (CF1) is composed of five kinds of subunits with a stoichiometry of alpha 3 beta 3 gamma delta epsilon. Reconstitution of a catalytically active alpha 3 beta 3 gamma core from urea-denatured subunits at a physiological pH is reported here. A restoration of approximately 90% of the CF1 ATPase activity has been observed. The reconstitution was achieved by using subunits overexpressed in Escherichia coli, purified, and combined in the presence of MgATP, K+, and a mixture of several chloroplast molecular chaperones at pH 7.5. The combination of chaperonin 60 and chaperonin 24 failed to reconstitute the active CF1 core, as did the GroEL/GroES pair (E. coli chaperonin 60/10 homologues). Characteristics of the reconstituted ATPase were very close to those of the native complex, including methanol-reversible inhibition by the purified epsilon subunit of CF1 and sensitivity to inhibition by azide and by tentoxin. In reconstitution with a mixture of tentoxin-resistant and -sensitive beta subunits, the extent of inhibition by tentoxin depended on the proportion of sensitive subunits in the reconstitution mixture. Finally, a model for the assembly of the CF1 core alpha 3 beta 3 gamma structure is proposed.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=45258Documentos Relacionados
- Partial deglycosylation of chloroplast coupling factor 1 (CF1) prevents the reconstitution of photophosphorylation.
- The maize chloroplast genes for the beta and epsilon subunits of the photosynthetic coupling factor CF1 are fused.
- Isolation and characterization of a Chlamydomonas reinhardtii mutant lacking the gamma-subunit of chloroplast coupling factor 1 (CF1).
- Molecular Evolution and Nucleotide Sequences of the Maize Plastid Genes for the α Subunit of Cf1 (atpA) and the Proteolipid Subunit of Cf0 (atpH)
- Reconstitution of mononucleosomes: characterization of distinct particles that differ in the position of the histone core.