A pseudoknot in the 3' non-core region of the glmS ribozyme enhances self-cleavage activity.

Published

Journal Article

The recently described glmS ribozyme is a self-cleaving RNA sequence found in the 5' noncoding region of the transcript of the gene for glucosamine-6-phosphate (GlcN6P) synthase in many Gram-positive bacteria. This ribozyme is associated with the GlcN6P riboswitch, and ribozyme activity in response to binding of the metabolite, GlcN6P, is proposed to effect levels of gene expression. The previously defined core sequence of the GlcN6P-dependent ribozyme contained fewer than 80 nt of contiguous sequence, but a sequence containing conserved secondary structural features and encompassing the core was twice as long. Structural elements outside of the ribozyme core could contribute to ribozyme activity or participate in gene regulation as part of the expression platform or both. Here, a 174-nt transcript containing the Bacillus anthracis glmS ribozyme was used to examine the contribution of part of the non-core sequence to in vitro cleavage activity. The loop portion of hairpin loop 3, located just 3' of the ribozyme core, can potentially pair with a sequence approximately 80 nt downstream to form a pseudoknot tertiary interaction. Disruptive and compensatory mutations in the two duplex regions of the pseudoknot had effects on in vitro cleavage rates that support a role for the pseudoknot in enhanced ribozyme activity. Cleavage activity became less sensitive to disruptive mutations in the pseudoknot as MgCl(2) concentrations were raised from 2.5 to 10 mM, suggesting that one role of the pseudoknot could be to help stabilize the core structure.

Full Text

Duke Authors

Cited Authors

  • Wilkinson, SR; Been, MD

Published Date

  • December 2005

Published In

Volume / Issue

  • 11 / 12

Start / End Page

  • 1788 - 1794

PubMed ID

  • 16314452

Pubmed Central ID

  • 16314452

International Standard Serial Number (ISSN)

  • 1355-8382

Digital Object Identifier (DOI)

  • 10.1261/rna.2203605

Language

  • eng

Conference Location

  • United States