Inhibitory mechanism of reveromycin A at the tRNA binding site of a class I synthetase.

The polyketide natural product reveromycin A (RM-A) exhibits antifungal, anticancer, anti-bone metastasis, anti-periodontitis and anti-osteoporosis activities by selectively inhibiting eukaryotic cytoplasmic isoleucyl-tRNA synthetase (IleRS). Herein, a co-crystal structure suggests that the RM-A molecule occupies the substrate tRNAIle binding site of Saccharomyces cerevisiae IleRS (ScIleRS), by partially mimicking the binding of tRNAIle. RM-A binding is facilitated by the copurified intermediate product isoleucyl-adenylate (Ile-AMP). The binding assays confirm that RM-A competes with tRNAIle while binding synergistically with L-isoleucine or intermediate analogue Ile-AMS to the aminoacylation pocket of ScIleRS. This study highlights that the vast tRNA binding site of the Rossmann-fold catalytic domain of class I aminoacyl-tRNA synthetases could be targeted by a small molecule. This finding will inform future rational drug design.

Aminoacyl-tRNA synthetases in Charcot-Marie-Tooth disease: A gain or a loss?

Charcot-Marie-Tooth disease (CMT) is one of the most common inherited neurodegenerative disorders with an increasing number of CMT-associated variants identified as causative factors, however, there has been no effective therapy for CMT to date. Aminoacyl-tRNA synthetases (aaRS) are essential enzymes in translation by charging amino acids onto their cognate tRNAs during protein synthesis. Dominant monoallelic variants of aaRSs have been largely implicated in CMT. Some aaRSs variants affect enzymatic activity, demonstrating a loss-of-function property. In contrast, loss of aminoacylation activity is neither necessary nor sufficient for some aaRSs variants to cause CMT. Instead, accumulating evidence from CMT patient samples, animal genetic studies or protein conformational analysis has pinpointed toxic gain-of-function of aaRSs variants in CMT, suggesting complicated mechanisms underlying the pathogenesis of CMT. In this review, we summarize the latest advances in studies on CMT-linked aaRSs, with a particular focus on their functions. The current challenges, future direction and the promising candidates for potential treatment of CMT are also discussed.

Aminoacyl tRNA synthetases as potential drug targets of the Panthera pathogen Babesia.

BACKGROUND: A century ago, pantheras were abundant across Asia. Illegal hunting and trading along with loss of habitat have resulted in the designation of Panthera as a genus of endangered species. In addition to the onslaught from humans, pantheras are also susceptible to outbreaks of several infectious diseases, including babesiosis. The latter is a hemoprotozoan disease whose causative agents are the eukaryotic parasites of the apicomplexan genus Babesia. Babesiosis affects a varied range of animals including humans (Homo sapiens), bovines (e.g. Bos taurus), pantheras (e.g. Panthera tigris, P. leo, P. pardus) and equines. Babesia spp. are transmitted by the tick vector Ixodes scapularis or ticks of domestic animals, namely Rhipicephalus (Boophilus) microplus and R. (B.) decoloratus. At the level of protein translation within these organisms, the conserved aminoacyl tRNA synthetase (aaRS) family offers an opportunity to identify the sequence and structural differences in the host (Panthera) and parasites (Babesia spp.) in order to exploit these for drug targeting Babesia spp. METHODS: Using computational tools we investigated the genomes of Babesia spp. and Panthera tigris so as to annotate their aaRSs. The sequences were analysed and their subcellular localizations were predicted using Target P1.1, SignalP 3.0, TMHMM v.2.0 and Deeploc 1.0 web servers. Structure-based analysis of the aaRSs from P. tigris and its protozoan pathogens Babesia spp. was performed using Phyre2 and chimera. RESULTS: We identified 33 (B. bovis), 34 (B. microti), 33 (B. bigemina) and 33 (P. tigris) aaRSs in these respective organisms. Poor sequence identity (~ 20-50%) between aaRSs from Babesia spp. and P. tigris was observed and this merits future experiments to validate new drug targets against Babesia spp. CONCLUSIONS: Overall this work provides a foundation for experimental investigation of druggable aaRSs from Babesia sp. in an effort to control Babesiosis in Panthera.

Possible therapy for ALS based on the cyanobacteria/BMAA hypothesis.

Although the cyanobacteria/BMAA hypothesis of the cause of ALS and other age-related neurodegenerative diseases remains to be proven, it is not too early to ask whether treatment would be possible if the hypothesis were correct. This paper reviews the possible ways that chronic BMAA neurotoxicity could be prevented or treated.

5-Fluorouracil modulates aminoacylation of rat liver tRNA.

Among the different classes of RNA, the effect of 5-fluorouracil (FUra), an anticancer drug, has been studied extensively on tRNA function in E. coli, but to a limited extent in eukaryotes, with specific reference to the mammalian system. Here we compared the aminoacylation function of rat liver tRNA substituted with FUra. Three hours after a single i.p. injection of 50, 250 or 500 mg/kg body wt. of FUra, total tRNAFUra50, tRNAFUra250, and tRNAFUra500, respectively were isolated from the livers of 2-3 month old male Wistar rats. The activity of tRNAFUra was compared with normal tRNA (tRNAN) isolated from saline-treated controls. tRNAFUra50 accepted [14C]-labeled total and five individual amino acids (lysine, aspartic acid, methionine, tryptophan, and serine) at a significantly higher rate compared to tRNAN. On contrary, tRNAFUra250 & 500 displayed a dose-dependent inhibition in aminoacylation with total amino acids, lysine, and methionine. Acceptance of leucine was inhibited by tRNAFUra in a dose-dependent way. Overall, the amino acid acceptance was variable among the three populations of rat liver tRNAFUra isolated with varying doses of FUra and the possible reasons for the altered function of tRNA are discussed.

Reaction of anti-OJ autoantibodies with components of the multi-enzyme complex of aminoacyl-tRNA synthetases in addition to isoleucyl-tRNA synthetase.

Autoantibodies to five aminoacyl-tRNA synthetases have been reported, and all have been associated with a syndrome of myositis and interstitial lung disease. Four of these synthetases exist free in the cytoplasm, but the fifth, isoleucyl-tRNA synthetase (recognized by anti-OJ autoantibodies), is a component of the multi-enzyme complex containing at least seven synthetases. In an effort to better understand the origins of these antibodies, we examined sera from 11 patients with anti-OJ autoantibodies for evidence of reaction with other components of the complex. All sera showed a characteristic pattern of 10 proteins bands by immunoprecipitation from HeLa cell extract. 10 of 11 sera significantly inhibited isoleucyl-tRNA synthetase enzyme activity. Serum and IgG from four patients also inhibited leucyl-tRNA synthetase activity, and serum and IgG from two inhibited lysyl-tRNA synthetase. Immunoblotting experiments supported reaction of the two sera with lysyl-tRNA synthetase, and revealed additional reactivity of three sera with a 160-kD component believed to be glutaminyl-tRNA synthetase. Despite reaction of some sera with additional synthetases, the immunoprecipitated tRNA appeared the same with all sera, and functioned as tRNA(ile). While reaction with more than one synthetase was seen with some anti-OJ sera, all synthetases targeted by anti-OJ sera were components of the complex, rather than unassociated synthetases. These findings suggest that an initial autoantibody response against isoleucyl-tRNA synthetase was followed by extension to involve other components of the synthetase complex. These observations may have implications for understanding the generation of antisynthetase autoantibodies.

[The identification of the sites of tRNA(Ser)(GCU) interaction in bovine liver with homologous aminoacyl-tRNA-synthetase by the chemical modification method]. / Vyznachennia dilianok vzaiemodiï tRNK(Ser)(GCU) pechinky byka iz homol'ohichnoiu aminoatsyl-tRNK-syntetazoiu metodom khemichnoï modyfikatsiï.

Interaction of the bovine liver tRNA(GCUSer) having a long variable loop, with the cognate aminoacyl-tRNA synthetase has been studied by alkylation with ethylnitrosourea. It was shown that seryl-tRNA synthetase protects 3'-phosphates of nucleotides 12, 13 in D-stem and 45-47-, 47 G.-, 47 H-variable stem of tRNA(GCUreS) from alkylation. An anticodon loop of tRNA(GCUSer) did not interact with seryl-tRNA synthetase.