Dataset Information

Cobalamin riboswitches exhibit a broad range of ability to discriminate between methylcobalamin and adenosylcobalamin.

ABSTRACT: Riboswitches are a widely distributed class of regulatory RNAs in bacteria that modulate gene expression via small-molecule-induced conformational changes. Generally, these RNA elements are grouped into classes based upon conserved primary and secondary structure and their cognate effector molecule. Although this approach has been very successful in identifying new riboswitch families and defining their distributions, small sequence differences between structurally related RNAs can alter their ligand selectivity and regulatory behavior. Herein, we use a structure-based mutagenic approach to demonstrate that cobalamin riboswitches have a broad spectrum of preference for the two biological forms of cobalamin in vitro using isothermal titration calorimetry. This selectivity is primarily mediated by the interaction between a peripheral element of the RNA that forms a T-loop module and a subset of nucleotides in the cobalamin-binding pocket. Cell-based fluorescence reporter assays in Escherichia coli revealed that mutations that switch effector preference in vitro lead to differential regulatory responses in a biological context. These data demonstrate that a more comprehensive analysis of representative sequences of both previously and newly discovered classes of riboswitches might reveal subgroups of RNAs that respond to different effectors. Furthermore, this study demonstrates a second distinct means by which tertiary structural interactions in cobalamin riboswitches dictate ligand selectivity.

SUBMITTER: Polaski JT

PROVIDER: S-EPMC5512062 | biostudies-literature | 2017 Jul

REPOSITORIES: biostudies-literature

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Cobalamin riboswitches exhibit a broad range of ability to discriminate between methylcobalamin and adenosylcobalamin.

Polaski Jacob T JT Webster Samantha M SM Johnson James E JE Batey Robert T RT

The Journal of biological chemistry 20170508 28

Riboswitches are a widely distributed class of regulatory RNAs in bacteria that modulate gene expression via small-molecule-induced conformational changes. Generally, these RNA elements are grouped into classes based upon conserved primary and secondary structure and their cognate effector molecule. Although this approach has been very successful in identifying new riboswitch families and defining their distributions, small sequence differences between structurally related RNAs can alter their l ...[more]

PMID: 28483920

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Project description:In bacteria, many essential metabolic processes are controlled by riboswitches, gene regulatory RNAs that directly bind and detect metabolites. Highly specific effector binding enables riboswitches to respond to a single biologically relevant metabolite. Cobalamin riboswitches are a potential exception because over a dozen chemically similar but functionally distinct cobalamin variants (corrinoid cofactors) exist in nature. Here, we measured cobalamin riboswitch activity in vivo using a Bacillus subtilis fluorescent reporter system and found, among 38 tested riboswitches, a subset responded to corrinoids promiscuously, while others were semiselective. Analyses of chimeric riboswitches and structural models indicate, unlike other riboswitch classes, cobalamin riboswitches indirectly differentiate among corrinoids by sensing differences in their structural conformation. This regulatory strategy aligns riboswitch-corrinoid specificity with cellular corrinoid requirements in a B. subtilis model. Thus, bacteria can employ broadly sensitive riboswitches to cope with the chemical diversity of essential metabolites. IMPORTANCE Some bacterial mRNAs contain a region called a riboswitch which controls gene expression by binding to a metabolite in the cell. Typically, riboswitches sense and respond to a limited range of cellular metabolites, often just one type. In this work, we found the cobalamin (vitamin B12) riboswitch class is an exception, capable of sensing and responding to multiple variants of B12-collectively called corrinoids. We found cobalamin riboswitches vary in corrinoid specificity with some riboswitches responding to each of the corrinoids we tested, while others responding only to a subset of corrinoids. Our results suggest the latter class of riboswitches sense intrinsic conformational differences among corrinoids in order to support the corrinoid-specific needs of the cell. These findings provide insight into how bacteria sense and respond to an exceptionally diverse, often essential set of enzyme cofactors.

Dataset Information

Cobalamin riboswitches exhibit a broad range of ability to discriminate between methylcobalamin and adenosylcobalamin.

Publications

Cobalamin riboswitches exhibit a broad range of ability to discriminate between methylcobalamin and adenosylcobalamin.

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