Ribozymes

Ribozymes

There was a time when proteins were considered the only biological macromolecules capable of catalysis. The discovery of the catalytic activity of RNA has thus had a profound impact on the way biochemists think. A few enzymes with RNA components had been discovered, such as telomerase and RNase P, an enzyme that cleaves extra nucleotides off the 5' ends of tRNA precursors. It was later shown that the RNA portion of RNase P has the catalytic activity. The field of catalytic RNA (ribozymes) was launched in earnest by the discovery of RNA that catalyzes its own self-splicing. It is easy to see a connection between this process and the splicing of mRNA by snRNPs. More recently, it has been shown that RNAs can catalyze reactions involved in protein synthesis. The catalytic efficiency of catalytic RNAs is less than that of protein enzymes, and the catalytic efficiency of currently existing RNA systems is greatly enhanced by the presence of protein subunits in addition to the RNA. Recall that many important coenzymes include an adenosine phosphate moiety in their structure. Compounds of such central importance in metabolism must be of ancient origin, another piece of evidence in support of the idea of an RNA-based world, where RNA was the original genetic molecule and the original catalytic one as well.

What are the characteristics of a ribozyme?

Several groups of ribozymes are known to exist. In Group I ribozymes, there is a requirement for an external guanosine, which becomes covalently bonded to the splice site in the course of excision. An example is the self-splicing that takes place in pre-rRNA of the ciliate protist Tetrahymena (Figure 11.38). The transesterification (of phosphoric acid esters) that takes place here releases one end of the intron. The free 3'-OH end of the exon attacks the 5' end of the other exon, splicing the two exons and releasing the intron. The free 3'-OH end of the intron then attacks a nucleotide 15 residues from the 5' end, cyclizing the intron and releasing a 5' terminal sequence. The precision of this sequence of reactions depends on the folded conformation of the RNA, which remains internally hydrogen bonded throughout the process. In vitro, this catalytic RNA can act many times, being regenerated in the usual way for a true catalyst. In vivo, however, it appears to act only once by splicing itself out. Group II ribozymes display a lariat mechanism of operation that was facilitated by snRNPs. There is no requirement for an external nucleotide; the 2'-OH of an internal adenosine attacks the phosphate at the 5' splice site. Clearly, DNA cannot self-splice in this fashion because it does not have a 2'-OH.

The folding of the RNA is crucial to its catalytic activity, as is the case with protein catalysts. A divalent cation (Mg²⁺ or Mn²⁺) is required; it is quite likely that metal ions stabilize the folded structure by neutralizing some of the negative charges on the phosphate groups of the RNA. A divalent cation is essential for the functioning of the smallest ribozymes known, the hammerhead ribozymes, which can be catalytically active with as few as 43 nucleotides. (The name comes from the fact that their structures resemble the head of a hammer when shown in conventional representations of hydrogen-bonded secondary structure.) The folding of RNA is such that large-scale conformational changes can take place with great precision. Similar large-scale changes take place in the ribosome in protein synthesis and in the spliceosome in the processing of mRNA. Note that they remain RNA machines when proteins have taken over much of the catalytic functioning of the cell. The ability of RNA to undergo the requisite large-scale conformational changes may well play a role in the process. A recently proposed clinical application of ribozymes has been suggested. If a ribozyme can be devised that can cleave the RNA genome of HIV, the virus that causes AIDS, it will be a great step forward in the treatment of this disease. Research on this topic is in progress in several laboratories.

Summary

Proteins are not the only biological molecules with catalytic properties. Some RNAs, called ribozymes, also catalyze certain reactions.

Group I ribozymes require an external guanosine for reactivity. Group II ribozymes do not have this requirement. They carry out catalysis via a lariat mechanism.