Small self-cleaving ribozymes

Abstract

The hammerhead, hairpin, hepatitis delta virus (HDV), Varkud Satellite (VS), and glmS ribozymes catalyze sequence-specific intramolecular cleavage of RNA. They range between 50 and 150 nucleotides in length, and are known as the "small self-cleaving ribozymes." Except for the glmS ribozyme that functions as a riboswitch in Gram-positive bacteria, they were originally discovered as domains of satellite RNAs. However, recent studies show that several of them are broadly distributed in genomes of organisms from many phyla. Each of these ribozymes has a unique overall architecture and active site organization. Crystal structures have revealed how RNA active sites can bind preferentially to the transition state of a reaction, whereas mechanistic studies have shown that nucleobases can efficiently perform general acid-base and electrostatic catalysis. This versatility explains the abundance of ribozymes in contemporary organisms and also supports a role for catalytic RNAs early in evolution.

Figure 1.

Internal transesterification reaction catalyzed by the hammerhead, hairpin, HDV, VS, and glmS ribozymes. The concerted cleavage reaction proceeds without intermediates. The hammerhead, hairpin, and VS ribozymes also catalyze the ligation reaction.

Figure 2.

RNase A active site structure and catalytic mechanism. The degree of impairment resulting from site-directed mutations of the catalytic residues (Raines 1998) is indicated in parentheses.

Figure 3.

Cartoon representations of the overall structures of four self-cleaving ribozymes. (A) The hairpin ribozyme, (B) the hammerhead ribozyme, (C) the glmS ribozyme-riboswitch, and (D) the hepatitis delta virus (HDV) ribozyme. Residues implicated in general acid and base catalysis in the cleavage reaction are green and red, respectively. The scissile phosphate and nucleophilic 2′-O atoms are magenta, flanked by substrate residues shown in light blue.

Figure 4.

Active sites of the (A) hairpin, (B) hammerhead, (C) glmS, and (D) HDV ribozymes color-coding as in Figure 3. The moieties thought to contribute to catalysis are labeled.

Figure 5.

Rolling-circle replication of satellite RNAs. Single-stranded covalently closed circles of sense (or genomic) RNA (red circle) are replicated by an RNA polymerase of the host, which transcribes processively, generating a linear multimeric concatamer (blue) that is complementary to the monomeric satellite RNA sense-strand. The multimeric complementary copy is then cleaved in to monomeric fragments, and these antigenomic RNAs circularize to produce templates for the second half of the replicative cycle, in which copies of the genomic sense strand are ultimately produced. The cleavage and ligation sites are often comprised of conserved ribozyme sequences that catalyze both phosphodiester bond cleavage and, in the case of circularization of the monomeric fragments, phosphodiester bond ligation. In the case of the satellite RNA of tobacco ringspot virus, the cleavage site in the sense strand is a hammerhead ribozyme, and that in the antisense strand is the hairpin ribozyme. HDV ribozyme sequences are present in both the sense and antisense strands of HDV.