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A study was conducted to study the influence of the 3'-CCA end of transfer RNA (tRNA) in the binding of catalytic magnesium ions by ribonuclease P. The cleavage reaction rate was represented as a function of mutant pre-tRNAs. The secondary structure of in vitro transcript pre-tRNA of Bacillus subtilis was also determined in the study. Results showed that the 3'-CCA and the 3'-proximal C residue support an important role in catalysis through the binding of catalytic Mg2+ ions.

A set of new ribozymes, where the RNaseP RNA is linked to a pre-tRNA substrate covalently, is designed to manipulate and investigate the function of RNaseP. Mature 5' end of tRNAs are produced by the RNaseP by the removal of 5'leader sequences from the pre-tRNAs. This reaction is catalyzed by the presence of RNA subunits, which function as ribozymes. The RNaseP exhibits sequence-specific cleavage by the Endo.P, and finds application in gene therapy.

Kinetic analysis revealed that the ribozyme ribonuclease (RNase) P catalytic mechanism has a cooperative reliance upon positively charged Magnesium concentration because a minimum of 3 positively charged Magnesium ions are needed for optimal activity, hinting at a multiple ion mechanism. The study proves the existence of a hyperbolic (saturable) reliance on hydroxide concentration, with half-maximal rate attained at pH 8.0-8.5.

The role of the four variable elements in Escherichia coli ribonuclease P (RNase P) was examined. The RNase P activity in vitro was not affected by the removal of one of the elements and the correct folding of the RNA is determined by two other elements. In addition, the structural stability and substrate binding affinity of the enzyme is influenced by long-range structural interactions participated in by a fourth element. It is demonstrated that significant functions of phylogenetically variable structures are not predicted by simple phylogenetic secondary structure comparison.