Molecular and cellular studies of tryptophanyl‐tRNA synthetases using monoclonal antibodies

Abstract

The content of Trp-tRNA synthetase in pancreas and liver of cattle, sheep, swine, rat, rabbit and man was assayed by direct radioimmunoblotting with a ' 2sI-labelled monoclonal antibody A m l , specifically interacting with any eukaryotic Trp-tRNA synthetase. Its content in the organs studied, with the exccption of bovine and sheep pancreas, was found to be 0.002-0.012% of total proteins. The enzyme content in bovine pancreas was about 0.2% of total proteins, i.e. 70 times higher than in bovine liver; similar correlations werc found for sheep. The Trp-tRNA synthetase levels in each organ varied from animal to animal of the same species by not more than a factor of four; these individual variations cannot affect the conclusion about the profound differences in the levels of the enzyme in pancreases of Rurninantia and of the other mammalians.As shown by indirect iminunofluorescence technique, bovine Trp-tRNA synthetase is mainly located in the exocrine part of the pancreas. Moreover, the immunoreactive material is detectable also in bovine (not human) pancreatic juice. The abnormally high Trp-tRNA synthetase content in the ruminant pancreas may be connected with unknown function(s) of this protein somehow related to the peculiarities of digestion of these mammals.Aminoacyl-tRNA synthetases play a key role in protein synthcsis by catalysing the highly specific formation of aminoacyl-tRNAs [l, 21. Apart from the aminoacylation reaction some synthetases catalyse the synthesis of a dinucleotide, adenosine( 5')-tetraphospho(5')adenosine [3], which plays an unknown role in cellular metabolism. Different organs and tissues are known to vary in their level of aminoacyl-tRNA synthetase activities [4], which may be due to the intensity of protein synthesis and the amino acid composition of proteins being synthesized [5], as well as to the hypothetical regulatory functions of these enzymes. The extremely high level of TrptRNA synthetase activity was revealed in bovine pancreatic tissue long ago [6].The application of immunochemical techniques allows one to directly measure the protein content, which does not always coincide with the enzyme activity level. The content of TrptRNA synthetase was assayed in different bovine organs using polyclonal antibodies against the bovine enzyme [7, 81. A very high Trp-tRNA synthetase content in the pancreas was revealed: 2-3% of the amount of soluble proteins. This content is two orders of magnitude higher than the enzyme level in several other bovine organs such as liver, kidney and heart [8], which suggests that, besides tRNA aminoacylation, TrpCorrcspondtnce to L. L. Kisselev, Institut Molekularnoj Biologii, Akadeniiya Nauk S.S.S.R., Vavilova ulitsa 32, Moskva, USSR 1 17984Abbreviations. RIPA buffer, 20 mM Tris/HCI. pH 7.5, 1 mM EDTA, IYu Nonidet P40, 1 % sodium deoxycholate, 0.1% SDS, 0.25 M NaCI.Enzyme. Tryptophanyl-tRNA synthetase (EC 6.1.1.2).tRNA synthetase of the bovine pancreas fulfills some additional, yet unknown, function(s). The goal of the present work was to establish wheth...

A human aminoacyl-tRNA synthetase as a regulator of angiogenesis

“…In bovine pancreas, fragments of TrpRS are present in pancreatic secretions (22,23), raising the possibility that proteolytic forms of TrpRS, in addition to the alternative spliced variant, might have biological activity. Because PMN elastase can release the active cytokines mini TyrRS and the endothelial monocyteactivating polypeptide II-like COOH domain from fulllength TyrRS (1), we examined PMN elastase cleavage of fulllength TrpRS.…”

“…Stimulation by IFN-␥ also induces production of the angiostatic chemokines IP-10 and MIG (19,20) and attenuates expression of the angiogenic chemokine IL-8 (21). Bovine full-length and truncated TrpRS, in which the extra NH 2 -terminal domain is deleted by proteolysis (5), are highly expressed in the pancreas and secreted into the pancreatic juice (22,23). The existence of various truncated forms of TrpRS as well as the close similarity between mammalian TyrRS and mammalian TrpRS raised the possibility that truncated TrpRSs have a function in addition to aminoacylation (5,24).…”

Immuno‐chemical non‐cross‐reactivity between eukaryotic and prokaryotic seryl‐tRNA synthetases

“…(2), bovine tyrosyl-tRNA synthetase (3), bovine tryptophanyl-tRNA synthetase (4). SerRS from Thermus thermophihts (5), lyzates of human K-562 cells (6), yeast (7) and E. coli (8).…”

“…The most characterized mammalian aminoacyl-tRNA synthetase by immunochemical methods is the tryptophanyl-tRNA synthetase from beef pancreas [5][6][7].…”

A Root‐Specific Iron‐Regulated Gene of Tomato Encodes a Lysyl‐tRNA‐Synthetase‐Like Protein

“…Different organs and tissues can vary considerably in the level of aminoacyl-tRNA synthetase activities. Thus, for instance, a tryptophanyl-tRNA synthetase with similarity to eukaryotic peptide-release factor (Garret et al, 1991;Lee et al, 1990) is strongly induced by interferon (Turpaev et al, 1996), and the amount of this enzyme found in pancreas greatly exceeds that necessary for protein biosynthesis (Sallafranque et al, 1986;Favorova et al, 1989).…”