RNA passes through the hole of the protein hexamer in the complex with the Escherichia coli Rho factor.
Escherichia coli transcription termination factor Rho is a ring-shaped hexameric protein that uses the energy derived from ATP hydrolysis to dissociate RNA transcripts from the ternary elongation complex. To test a current model for the interaction of Rho with RNA, three derivatives of Rho were made containing single cysteine residues and modified with a photo-activable cross-linker. The positions for the cysteines were: 1) in part of the primary RNA-binding site in the N terminus (Cys-82 Rho); 2) in a connecting polypeptide proposed to be on the outside of the hexamer (Cys-153 Rho); and 3) near the proposed secondary RNA-binding site in the ATP-binding domain (Cys-325 Rho). Results from the cross-linking of the modified Rho proteins to a series of lambda cro RNA derivatives showed that Cys-82 Rho formed cross-links with all transcripts containing the Rho utilization (rut) site, that Cys-325 Rho formed cross-links to transcripts that had the rut site and 10 or more residues 3' of the rut site, and that Cys-153 did not form cross-links with any of the transcripts. From a model of the quaternary structure of Rho, which is largely based on homology to the F(1)-ATPase, amino acid 82 is located near the top of the hexamer, and amino acid 325 is located on a solvent-accessible loop in the center of the hexamer. These data are consistent with binding of the rut region of RNA around the crown, with its 3'-segment passing through the center of the Rho hexamer.
Burgess
B R
BR
Department of Chemistry, Indiana University, Bloomington, Indiana 47405, USA.
Richardson
J P
JP
D000255:Adenosine Triphosphate / Q000378:metabolism; D001483:Base Sequence; D017931:DNA Primers; D004926:Escherichia coli / Q000378:metabolism*; D006868:Hydrolysis; D011487:Protein Conformation; D012329:RNA, Bacterial / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism*; D012234:Rho Factor / Q000737:chemistry / Q000378:metabolism*
D017931:DNA Primers; D012329:RNA, Bacterial; D012333:RNA, Messenger; D012234:Rho Factor; D000255:Adenosine Triphosphate