C-terminal Domain of Leucyl-tRNA Synthetase from Pathogenic Candida albicans Recognizes both tRNASer and tRNALeu

Quan-Quan Ji,Zhi-Peng Fang,Qing Ye,Zhi-Rong Ruan,Xiao-Long Zhou,En-Duo Wang

doi:10.1074/jbc.m115.699777

Abstract

Leucyl-tRNA synthetase (LeuRS) is a multidomain enzyme that catalyzes Leu-tRNA(Leu) formation and is classified into bacterial and archaeal/eukaryotic types with significant diversity in the C-terminal domain (CTD). CTDs of both bacterial and archaeal LeuRSs have been reported to recognize tRNA(Leu) through different modes of interaction. In the human pathogen Candida albicans, the cytoplasmic LeuRS (CaLeuRS) is distinguished by its capacity to recognize a uniquely evolved chimeric tRNA(Ser) (CatRNA(Ser)(CAG)) in addition to its cognate CatRNA(Leu), leading to CUG codon reassignment. Our previous study showed that eukaryotic but not archaeal LeuRSs recognize this peculiar tRNA(Ser), suggesting the significance of their highly divergent CTDs in tRNA(Ser) recognition. The results of this study provided the first evidence of the indispensable function of the CTD of eukaryotic LeuRS in recognizing non-cognate CatRNA(Ser) and cognate CatRNA(Leu). Three lysine residues were identified as involved in mediating enzyme-tRNA interaction in the leucylation process: mutation of all three sites totally ablated the leucylation activity. The importance of the three lysine residues was further verified by gel mobility shift assays and complementation of a yeast leuS gene knock-out strain.

Highlights

Aminoacyl-tRNA synthetases3 are a family of enzymes that catalyze aminoacyl-tRNA formation, playing a pivotal role in protein translation [1]
The CaCTD Is Indispensable for Leucylating Both CatRNASer(CAG) and CatRNALeu—Our previous study showed that only eukaryotic Leucyl-tRNA synthetase (LeuRS) (e.g. cytoplasmic LeuRS (CaLeuRS), Saccharomyces cerevisiae LeuRS (ScLeuRS), and Homo sapiens LeuRS (HsLeuRS)) could charge CatRNASer(CAG); archaeal LeuRS (e.g. Pyrococcus horikoshii LeuRS (PhLeuRS)) and bacterial/mitochondrial LeuRS (e.g. EcLeuRS and CamtLeuRS) could not [24]
The archaeal and eukaryotic LeuRSs appear to be derived from the same ancestor with the most divergence in the C-terminal domain (CTD), implying that this region may be critical for recognizing CatRNASer

Summary

Introduction

Aminoacyl-tRNA synthetases (aaRSs) are a family of enzymes that catalyze aminoacyl-tRNA formation, playing a pivotal role in protein translation [1]. Leucyl-tRNA synthetase (LeuRS), which is a class I aaRS, is further classified into bacterial and archaeal/eukaryotic types on the basis of CP1 domain insertion site and orientation (6 – 8). Both types of LeuRS possess a catalytic domain (for amino acid activation and tRNA charging), a CP1 domain (for editing), an ␣-helix bundle domain, and a C-terminal domain (CTD; for tRNA binding) [9]. The element in CaSerRS that recognizes tRNASer has been revealed Another critical aaRS involved in CUG decoding ambiguity, CaLeuRS, with 1,097 residues, contains a single CUG codon-encoded residue at position 919, located in its CTD (from Gly894 to Glu1097) [24]. Eukaryotic LeuRS and archaeal PhLeuRS only exhibit obvious divergence in their CTD, indicating the potential significance of the CTD of eukaryotic LeuRS in tRNASer(CAG) recognition

Results

Discussion

Conclusion