In:
RNA, Cold Spring Harbor Laboratory, Vol. 13, No. 4 ( 2007-04), p. 521-535
Abstract:
Bacterial ribonuclease P (RNase P) is a ribonucleoprotein complex composed of one catalytic RNA (PRNA) and one protein subunit (P protein) that together catalyze the 5′ maturation of precursor tRNA. High-resolution X-ray crystal structures of the individual P protein and PRNA components from several species have been determined, and structural models of the RNase P holoenzyme have been proposed. However, holoenzyme models have been limited by a lack of distance constraints between P protein and PRNA in the holoenzyme–substrate complex. Here, we report the results of extensive cross-linking and affinity cleavage experiments using single-cysteine P protein variants derivatized with either azidophenacyl bromide or 5-iodoacetamido-1,10- o -phenanthroline to determine distance constraints and to model the Bacillus subtilis holoenzyme–substrate complex. These data indicate that the evolutionarily conserved RNR motif of P protein is located near ( 〈 15 Å) the pre-tRNA cleavage site, the base of the pre-tRNA acceptor stem and helix P4 of PRNA, the putative active site of the enzyme. In addition, the metal binding loop and N-terminal region of the P protein are proximal to the P3 stem–loop of PRNA. Studies using heterologous holoenzymes composed of covalently modified B. subtilis P protein and Escherichia coli M1 RNA indicate that P protein binds similarly to both RNAs. Together, these data indicate that P protein is positioned close to the RNase P active site and may play a role in organizing the RNase P active site.
Type of Medium:
Online Resource
ISSN:
1355-8382
,
1469-9001
Language:
English
Publisher:
Cold Spring Harbor Laboratory
Publication Date:
2007
detail.hit.zdb_id:
1475737-0
SSG:
12
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