Plasmodium apicoplast tyrosyl-tRNA synthetase recognizes an unusual, simplified identity set in cognate tRNATyr

• Published in: PloS one 2018-12, Vol.13 (12), p.e0209805-e0209805
• Main Authors: Cela, Marta, Paulus, Caroline, Santos, Manuel A S, Moura, Gabriela R, Frugier, Magali, Rudinger-Thirion, Joëlle
• Format: Article
• Language: English
• Subjects: Amino acids; Amino Acyl-tRNA Synthetases - genetics; Amino Acyl-tRNA Synthetases - metabolism; Aminoacylation; Apicoplasts - enzymology; Apicoplasts - metabolism; Biology and life sciences; Biosynthesis; Cloning; E coli; Enzymes; Genes; Genomes; Humans; Life cycle engineering; Life cycles; Life Sciences; Malaria; Malaria, Falciparum - physiopathology; Physical Sciences; Plasmodium; Plasmodium falciparum; Plasmodium falciparum - enzymology; Plasmodium falciparum - metabolism; Plasmodium falciparum - pathogenicity; Protein synthesis; Proteins; Protozoan Proteins - genetics; Protozoan Proteins - metabolism; Recognition; Research and Analysis Methods; RNA polymerase; RNA, Transfer, Tyr - genetics; RNA, Transfer, Tyr - metabolism; Substrates; tRNA; tRNA Tyr; Tyrosine-tRNA ligase; Tyrosine-tRNA Ligase - genetics; Tyrosine-tRNA Ligase - metabolism; Vector-borne diseases
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0209805

The life cycle of Plasmodium falciparum, the agent responsible for malaria, depends on both cytosolic and apicoplast translation fidelity. Apicoplast aminoacyl-tRNA synthetases (aaRS) are bacterial-like enzymes devoted to organellar tRNA aminoacylation. They are all encoded by the nuclear genome and are translocated into the apicoplast only after cytosolic biosynthesis. Apicoplast aaRSs contain numerous idiosyncratic sequence insertions: An understanding of the roles of these insertions has remained elusive and they hinder efforts to heterologously overexpress these proteins. Moreover, the A/T rich content of the Plasmodium genome leads to A/U rich apicoplast tRNA substrates that display structural plasticity. Here, we focus on the P. falciparum apicoplast tyrosyl-tRNA synthetase (Pf-apiTyrRS) and its cognate tRNATyr substrate (Pf-apitRNATyr). Cloning and expression strategies used to obtain an active and functional recombinant Pf-apiTyrRS are reported. Functional analyses established that only three weak identity elements in the apitRNATyr promote specific recognition by the cognate Pf-apiTyrRS and that positive identity elements usually found in the tRNATyr acceptor stem are excluded from this set. This finding brings to light an unusual behavior for a tRNATyr aminoacylation system and suggests that Pf-apiTyrRS uses primarily negative recognition elements to direct tyrosylation specificity.