This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.The metabolic state of a cell typically affects gene expression through protein-mediated mechanisms of transcriptional or translational control. Yet recent studies have shown that messenger RNAs (mRNAs) can directly sense and respond to specific metabolites through intrinsic domains termed riboswitches. In the presence of metabolites, structural changes in riboswitch domains can result in either transcriptional termination or translational repression. Recently a novel catalytic riboswitch has been discovered that exerts genetic control through self-cleavage of the nascent RNA in response to cellular metabolite concentration. The metabolite-dependent ribozyme resides in the 5'-untranslated region of the glmS mRNA of numerous Gram-positive bacteria and it catalyzes an internal phosphoester transfer reaction that results in cleavage and inactivation of the mRNA. The ribozyme selectively recognizes and is 1000-fold activated by glucosamine-6-phosphate, the metabolic product of the GlmS enzyme, and an important component of bacterial cell walls. We are interested in understanding the molecular basis of ligand recognition and catalysis by the glmS riboswitch. To address this we measured the rate of self-cleavage of the ribozyme/riboswitch in response to a panel of related but distinct ligands, serinol, glucose and glucosamine, to name a few. We have determined that the ribozyme makes important contacts to the amine and phosphate in the natural ligand. We are interested in determining how these two functional groups are interacting with the full-length glmS ribozyme in order to fully understand its role in catalysis.
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