5.4.99.24	D139N	site-directed mutagenesis of the chimeric enzyme mutant RluCD, the mutant is catalytically inactive	748788	
5.4.99.24	additional information	a proteolytically derived fragment of the enzyme consisting of residues 89-319 retains catalytic activity	5857	
5.4.99.24	additional information	construction of a chimeric pseudouridine synthase (RluCD) containing the N-terminal S4 domain of enzyme RluC (EC 5.4.99.24) and the C-terminal catalytic domain of enzyme RluD (EC 5.4.99.23). The chimeric mutant is able to introduce excessive pseudouridines into rRNA at non-native positions. The chimeric enzyme RluCD is used as a tool to study an effect of over-modification of rRNA on the ribosome biogenesis. Excessive pseudouridylation of 23S rRNA reduces progression of ribosome assembly during early or middle stages. A modification interference approach identifies the sites in 23S rRNA whose modification prevents ribosome assembly. It is plausible that pseudouridines can cause RNA misfolding when present at non-native positions. RluCD isomerizes many uridines of rRNA in a non-specific manner. Induction of the RluCD at the exponential growth phase leads to severe inhibition of translation while transcription is only slightly affected	748788