Abstract
The active form of transcription termination factor rho from Escherichia coli is a homohexamer, but several studies suggest that the six subunits of the hexamer are not functionally identical. Rho has three tight and three weak ATP binding sites. Based on our findings, we propose that the tight nucleotide binding sites are noncatalytic and the weak sites are catalytic. In the presence of RNA, the rho-catalyzed ATPase rate is fast, close to 30 s-1. However, under these conditions the three tightly bound nucleotides dissociate from the rho hexamer at a slow rate of 0.02 s-1, indicating that the three tight nucleotide binding sites of rho do not participate in the fast ATPase turnover. These slowly exchanging nucleotide binding sites of rho are capable of hydrolyzing ATP, but the resulting products (ADP and Pi) bind tightly and dissociate from rho about 1500 times slower than the fast ATPase turnover. Both RNA and excess ATP in solution are necessary for stabilizing nucleotide binding at these sites. In the absence of RNA, or when solution ATP is hydrolyzed to ADP, a faster dissociation of nucleotides was observed. Based on these results, we propose that the rho hexamer is similar to the F1-ATPase and T7 DNA helicase-containing noncatalytic sites that do not participate in the fast ATPase turnover. We propose that the three tight sites on rho are the noncatalytic sites and the three weak sites are the catalytic sites.
Highlights
The transcription termination factor rho is an essential protein in Escherichia coli that is required for the release of certain nascent RNAs from the transcription complex [1,2,3,4]
Because rho protein is structurally similar to the F1-ATPase protein, we have investigated the possibility of noncatalytic sites on the rho hexamer
We propose that the three tight nucleotide binding sites on the rho hexamer are noncatalytic, serving a role similar to such sites of the F1-ATPase and T7 DNA helicase, and that the weak sites are the catalytic sites
Summary
The transcription termination factor rho is an essential protein in Escherichia coli that is required for the release of certain nascent RNAs from the transcription complex [1,2,3,4]. The rho protein contains an ATPase-dependent helicase activity that can separate the strands of an RNA-DNA duplex. A recent report has shown similarities among the ATPase mechanism of rho, the F1-ATPase, and T7 DNA helicase [23, 24] These observations support the proposal that the structure of the rho hexamer is likely to be similar to that of the F1-ATPase (14 –16). The three tight nucleotide binding sites on T7 DNA helicase hexamer were shown to be noncatalytic, with properties similar to such sites of the F1-ATPase protein. We propose that the three tight nucleotide binding sites on the rho hexamer are noncatalytic, serving a role similar to such sites of the F1-ATPase and T7 DNA helicase, and that the weak sites are the catalytic sites. ¶ Supported by Core Research Grant C2-10680651 from the Ministry of Education, Science, Culture and Sports of Japan
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
