## 4 Antibiotics Inhibiting Aminoacylation of tRNA
### Inhibition by Trapping tRNA in an \"Aminoacylation-Like\" Conformation  
Agrocin 84 (Figure 3) is a Trojan horse inhibitor that targets the pathogenic strains _Agrobacteriumumumefaciens_ and _Agrobacteriumum rhizogenes_ that in plants cause crown gall tumors and hairy root condition, respectively. The antibiotic is produced by another agrobacterial strain, _Agrobacteriumum radiobacter_ strain K84 and can act as a natural biocontrol agent and can prevent plant tumors in a number of agriculturally important species [99]. Tumorgenic _Agrobacterial_ strains contain a tumor-inducing or Ti-plasmid from which an oncogenic fragment of DNA called T-DNA is injected into wounded-plant cells through a molecular syringe and stably incorporated into the plant genome [100, 101]. The expression of the T-DNA genes leads to production of plant hormones as well as carbon-rich compounds called opines, which are secreted. The pathogenic agrobacteria in turn metabolize these giving a rational for the infection process [101, 102].  
Figure 3: Chemical structures of tRNA based inhibitors of aminoacyl-tRNA synthetases (AN2690 and agrocin 84/TM84) and ribosome (thermorubin and blasticidin S).  
Agrocin 84 acts as a molecular Trojan horse by mimicking a tumor-derived substrate called agrocinoping [103, 104] and so hijacks the _A. tumefaciens_' opine permease to gain access into the interior of the cell [105, 106, 107]. Once inside, agrocin 84 is processed into a toxic moiety called TM84 that inhibits LeuRS [108, 109] (Figure 3). TM84 is structurally similar to the leucyl-adenylate intermediate (Leu-AMP) that is formed in the first step of the LeuRS aminoacylation reaction. One of the key differences is that Leu-AMP possesses an unstable phosphoanhydride bond as opposed to TM84, which contains a stable _N_-acyl phosphoramidate bond [110]. Surprisingly, TM84 is a weak inhibitor of the amino acid activation reaction. This is unexpected as it suggests that TM84 does not behave as a standard aminoacyl adenylate analog discussed earlier. An explanation for this behavior has recently been found that demonstrates that another player is involved in the TM84 inhibition of LeuRS. Biochemical and biophysical analyses have revealed that tRNA\({}^{\text{Leu}}\) is required to form a tight binding complex of _A. tumefaciens_ LeuRS'tRNA\({}^{\text{Leu}}\).TM84 (IC\({}_{50}\sim\)1 nM, \(K_{\text{iapp}}\)= 0.3 nM) This was also confirmed by the X-ray crystal structure of the complex of _E. coli_ LeuRS'tRNA\({}^{\text{Leu}}\).TM84, which demonstrated that TM84 does indeed bind to the catalytic active site; But the CCA 3'-end of tRNA\({}^{\text{Leu}}\) is positioned into the enzyme active site, trapped into an \"aminoacylation-like\" conformation. Specifically, the terminal adenosine of the CCA 3'-end directly interacts with the TM84 toxin. Additionally, TM84 forms interactions with the highly conserved, mobile KMSKS loop of LeuRS, thereby allowing the loop to adopt a closed-conformation encapsulating TM84 and explaining the high affinity of the toxin in the presence of tRNA\({}^{\text{Leu}}\)[109]. The two novel tRNA-dependent inhibition mechanisms employed by TM84 and AN2690 highlighted here may well provide promising stepping-stones to development of tRNA-dependent inhibitors of the other amino acid specific aaRSs.