Probing Open Conformation of GroEL Rings by Cross-linking Reveals Single and Double Open Ring Structures of GroEL in ADP and ATP

AUTOR(ES)

Nojima, Tatsuya

FONTE

American Society for Biochemistry and Molecular Biology

RESUMO

Two heptamer rings of chaperonin GroEL undergo opening-closing conformational transition in the reaction cycle with the aid of GroES and ATP. We introduced Cys into the GroEL subunit at Ala-384 and Ser-509, which are very close between adjacent GroEL subunits in the open heptamer ring but far apart in the closed heptamer ring. The open ring-specific inter-subunit cross-linking between these Cys indicated that the number of rings in open conformation in GroEL was two in ATP (GroELOO), one in ADP (GroELO), and none in the absence of nucleotide. ADP showed an inhibitory effect on ATP-induced generation of GroELOO. The isolated GroELO and GroELOO, which lost any bound nucleotide, could bind GroES to form a bullet-shaped 1:1 GroEL-GroES complex and a football-shaped 1:2 GroEL-GroES complex, respectively, even without the addition of any nucleotide. Substrate protein was unable to form a stable complex with GroELOO and did not stimulate ATPase activity of GroEL. These results favor a model of the GroEL reaction cycle that includes a football complex as a critical intermediate.

Documentos Relacionados

• Cross-linking reconsidered: binding and cross-linking fields and the cellular response.
• Chemical cross-linking of Chlamydia trachomatis.
• Hybridization triggered cross-linking of deoxyoligonucleotides.
• NAD-dependent cross-linking of dinitrogenase reductase and dinitrogenase reductase ADP-ribosyltransferase from Rhodospirillum rubrum.
• Cross-linking da cornea: protocolo padrão