L-Amino Acids Determination by Isotope Dilution Technique in the tRNA Loading Test

Abstract

The transfer ribonucleic acids (tRNA) transfer amino acids to the peptide chain growing on the ribosome in protein biosynthesis. The amino acids are activated by enzymes known as amino-acyl-tRNA synthetases or amino-acyl-tRNA ligases. For each of the 20 amino acids that take part in protein biosynthesis, there is at least one specific tRNA and one synthetase. Both the tRNAs and the synthetases are highly specific with respect to the assigned amino acid. This property can be used for the quantitative determination of l -amino acids and has been exploited for some time for the determination of the acceptor activities of tRNA for amino acids and the activity of synthetases with the aid of radioactively labeled amino acids. This method can be recommended where one or a few amino acids are to be determined in several amino acid mixtures or where one amino acid is to be determined in many samples in a series of experiments. Manometric method is used for the determination of some l -amino acids, such as l -lysine, l -arginine, l -ornithine, l -tyrosine, l -histidine, l -glutamic acid, and l -aspartic acid. This method is applied in biochemistry, in the analysis of proteins, and in microbiology. Colorimetric method with fluorodinitrobenzene (FDNB) is used for the determination of l -lysine, l -arginine, l -histidine, l -ornithine, and l -tyrosine in plasma and urine, particularly for a series of determinations of a single amino acid, and for confirmation of the identity of an amino acid tentatively identified by other methods. The process can be easily extended to additional amino acids if corresponding enzymes are available. In the case of labeled amino acids in biological solutions, the carbon skeleton can be isolated by this method.