Competing addition and hydrolysis of the cytidylylcytidylyladenosine terminal residues of transfer ribonucleic acid isolated from the non-lactating bovine mammary gland

Abstract

1. The enzyme fraction obtained from the pH5 enzyme of non-lactating bovine mammary gland between 40 and 100% ammonium sulphate saturation markedly inhibited the AMP-incorporating activity of rat liver nucleotide-incorporating enzyme. This inhibitory effect has been attributed to high nuclease activity which can be partially removed by adsorption of the enzyme fraction on to calcium phosphate gel. 2. The degradation action of the calcium phosphate-purified enzyme is confined mainly to the terminal trinucleotide sequence -pCpCpA of tRNA, its effect being analogous to that of venom phosphodiesterase. This enzyme is heat labile and very readily loses its degradative activity. 3. Treatment of the enzyme fraction with Macaloid results in complete removal of the phosphodiesterase, leaving an enzyme capable of incorporating AMP into tRNA. 4. Transfer RNA extracted from non-lactating bovine mammary gland in the presence of polyvinyl sulphate and Macaloid is able to accept amino acids with an efficiency 30% of that shown by lactating bovine mammary-gland tRNA isolated under identical conditions.

Evidence for defective transfer ribonucleic acid in polymyopathic hamsters and its inhibitory effect on protein synthesis

“…It is known that addition of tRNA can lead to inhibition of the incorporation of amino acids in protein-synthesizing systems (Herrington & Hawtrey, 1970b). A significant feature here, however, is that tRNA isolated from the pH5 enzyme fraction from myopathic muscle had no stimulatory effect in both systems studied (Table 1), although it could be aminoacylated to a certain extent (Fig.…”

“…1). Herrington & Hawtrey (1970b) showed that bovine tRNA, lacking the terminal -pC-C-A-OH trinucleotide group, competitively inhibits the aminoacylation of rat liver tRNA. On the other hand, when the terminal -pC-C-A-OH group has been added the material stimulates the synthesis of rat liver aminoacyl-tRNA.…”

“…Activity of nucleotide-incorporating enzymes from myopathic and control hamster skeletal muscles tRNA nucleotidyltransferase was partially purified, by using pH5 enzyme fractions from myopathic and control muscles as starting material, and generally following the procedures of Herrington & Hawtrey (1970b). Twice-pyrophosphorolysed rat liver tRNA (lacking -pC-C-A-OH) was used as substrate.…”

“…132 (Cannon, 1966;Rosset & Monier, 1963), but the amino acid-acceptor capacity of tRNA depends on intact 3'-trinucleotide end-sequences. Thus removal of these terminal nucleotides, in part or whole, caused complete inability to accept amino acids (Hecht et al, 1958;Cannon, 1964Cannon, , 1965Herrington & Hawtrey, 1970b;Carre et al, 1970) or to bind to ribosomes (Cannon et al, 1963). As found by Rosset & Monier (1963) and by Cannon (1966) in Escherichia coli preparations, and by Herrington & Hawtrey (1970a,b) in bovine mammary glands, it is possible that ribonuclease II, or an exonuclease with a similar mechanism, hydrolytically removes the terminal adenylate group from tRNA molecules, causing inactivation.…”

Elevated levels of acceptor activity of hepatoma transfer ribonucleic acid

“…This enzyme has been extensively purified and studied in Escherichia coli (Carre et al, 1970), and it has been found in 55-85 % ammonium sulfate saturated fractions from rat liver (Herbert and Canellakis, 1963). The enzyme is active under the conditions of the aminoacylation assay (Herrington and Hawtrey, 1970) and the addition of CTP to reaction mixtures provides conditions suitable to the repair of tRNAs deficient at the CpCpA-OH terminus. The addition of CTP to reaction mixtures, and therefore the repair of deficient pCpCpA-OH termini, does not significantly increase the extent of liver phenylalanyl-tRNA formation, much less increase it to the level obtainable with hepatoma tRNA.…”