A protein was purified from rat liver which stimulated a number of liver aminoacyl-tRNA synthetases. This stimulatory factor was identical with the "tRNA activator" of Dickman & Boll [(1976) Biochemistry 15, 3925] in its mechanism of action and chemical properties, although it was considerably more purified. The two preparations stimulated synthetases by virtue of their pyrophosphatase activity which destroyed the potent inhibitor, PPi, that was present in the reaction mixtures. This PPi was either generated during the reaction or was introduced by contamination of the tRNA or ATP preparations. The degree of inhibition of PPi was strongly influenced by assay conditions, being most effective at low amino acid concentrations, at low pH, and in the presence of heterologous tRNAs. By use of certain assay conditions, PPi concentrations as low as 2 μM could inhibit some synthetases close to 50%. The pitfalls associated with some assay conditions commonly used for aminoacyl-tRNA synthetases are discussed. These studies raise questions about the physiological significance of many previously described aminoacyl-tRNA synthetase stimulatory factors.
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