Let us make an in-depth study of the ribozyme. After reading this article you will learn about: 1. Self Splicing Process or Autosplicing 2. Role of Ribozyme in Protein Synthesis and 3. Hammerhead Ribozyme.
Self Splicing Process or Autosplicing:
Like protein enzymes some RNA molecules function as enzymes. Like protein enzymes some RNA sequences form complex tertiary structures and play the role of biological catalysts. Such RNA enzymes are known as ribozymes. Like enzymes they have an active site, a binding site for a substrate and a binding site for a co-factor such as metal ion.
Ribozymes have been extensively studied in rRNA of a ciliate protozoan Tetrahymena thermophila and are also found in fungal mitochondria, in phage T4 and E. coli bacteria.
Group 1 introns have an intrinsic ability to splice themselves which is called self splicing or autosplicing. Here no enzyme is required and RNA sequence functions as its own ribozyme and behaves like an endonuclease. In rRNA of Tetrahymena the splicing process involves binding of a single guanosine (G) to a site within the intron which is to be spliced out.
The splicing involves two transesterification reactions. First a free guanosine nucleotide attacks the phosphodiester bond at the 5′ splice site of the intron cleaving the 5′ end of intron. Second trans-esterification reaction splices the phosphodiester bond at the 3′-splice site of next exon causing cleavage. This causes the 3′ end of exon 1 to form a phosphodiester bond with 5′ end of exon 2, thus joining them. The intron is released in a linear form, which is subsequently degraded.
It this way the RNA molecule functions as its own ribozyme as it splices itself without the help of any protein enzyme.
Another ribozyme is ribonuclease P (RNAse P) which is a ribonuclease involved in the processing of pre-tRNA. The RNAse P is an ribozyme composed of one RNA component and one protein component. Its RNA component is a ribozyme which catalyzes the processing of precursors tRNA.
Precursor tRNA is a very long molecule. During processing its large portion is removed and degraded. Precursor tRNA is cleaved at both 5′-end and 3′-end. In E. coli the tRNA which carries amino acid tyrosine (tRNAlyr) has been extensively studied. The cleavage occurs differently at 5′-end and 3′-end. At the 5′-end the precursor tRNA is recognized by an enzyme known as RNAse P which is a ribozyme. This ribozyme cleaves a portion of pre-tRNA so that correct 5′-end of mature tRNA is produced. This proves that all enzymes are not proteins.
Role of Ribozyme in Protein Synthesis:
In protein synthesis, during elongation phase, the amino acids are added one-by-one. During elongation phase, a tRNA brings a new amino acid to the ribosome according to the codons of mRNA. Each new amino acid is then attached to the end of the growing polypeptide chain. During chain elongation, a peptide bond is formed between the amino acid at “A” site and growing polypeptide chain, which is transferred from “P” site to “A” site. The transfer is known as peptidyl transfer reaction.
This transfer involves formation of peptide bond between amino acid at the “A” site and polypeptide chain, thus lengthening the chain by one amino acid. It was discovered that the peptidy transferase.
Which catalizes the peptide bond formation between successive amino acids consists of several proteins and a 23S rRNA molecule present in the ribosome. This 23S rRNA is a ribozyme and is responsible for catalyzing peptide bond formation between successive amino acid.
Hammerhead Ribozyme:
Hammerhead ribozyme is another small RNA which performs self cleavage reaction. It is found in viroids, which have RNA as genetic material and they infect the plants. When viroid replicates by rolling circle, it produces a continuous RNA chain, which consists of multiple copies of RNA. This continuous chain RNA undergoes cleavage to form single viroids or monomers. The self-cleaving RNA sequence at the junction of monomers is called hammerhead because of the shape of its secondary structure.
It consists of three complementary base paired stems. These are stem I, stem II and stem III surrounding a core of non-complementary nucleotides. This core lies at their junction and undergoes self-cleavage reaction.
The catalytic reaction centre lies between stem 2/3 and stem I. This centre contains a magnesium ion that initiates hydrolysis reaction. The reaction breaks the RNA chain producing a 2′, 3′ cyclic phosphate and a free 5′-hydroxyl end.
Like antisense RNA, which is used to silence the genes, rihozyme is also used to turn off specific genes.