Leucyl-tRNA Synthetase (Archaeal-like)

The archaeal-like leucyl-tRNA synthetase (LeuRS-A) is an enzyme that plays a crucial role in protein synthesis by catalyzing the attachment of the amino acid leucine to its cognate tRNA: $ \text{Leu} + \text{tRNA}^\text{Leu} + \text{ATP} \xrightarrow{\text{LeuRS-A}} \text{Leu-tRNA}^\text{Leu} + \text{AMP} + \text{PP}_i $ LeuRS-A is found in most archaea, and differs from the bacterial-like form [LeuRS-B](/class1/leu1), which is found in most bacteria and organelles. Either type may reside in a eukaryotic cytosol. The two forms are characterized by their abilities to charge tRNAs corresponding to six different codons. In the standard genetic code, [ArgRS](/class1/arg) and [SerRS](/class1/ser1) are the only other AARS which decode as many as six codons. The structures of LeuRS-A and -B are distinguished by i) the placement of the editing domain within their primary sequence (Fukunaga et al. 2005), ii) the C-terminal domain which recognises tRNA (Tukalo et al. 2005), and iii) the two LeuRS-A insertion modules depicted below, also known as the archaeal leucyl-specific domains 1 and 2 (Fukunaga et al. 2005). LeuRS-A and -B alike are closely related to the [IleRS](/class1/ile), [ValRS](/class1/val), and [MetRS](/class1/met) families, which comprise subclass Ia (Perona and Hadd 2012, Gomez and Ibba, 2020). Members of subclass Ia are characterized by their hydrophobic amino acid substrates, as well as the connecting peptide 2 (CP2) and zinc finger (ZF) insertions, depicted below. CP2 is a 30-40 amino acid two-helix bundle on the surface of the catalytic domain (Starzyk et al. 1987) and appears to be essential for amino acid activation (Zhou et al. 2008). Upstream from this resides a cysteine-rich zinc finger, 20-40 amino acids in length, also essential for effective aminoacylation (Nureki et al. 1993, Sugiura et al. 2000). However, zinc binding activity appears to be absent from this region of the protein for certain members of LeuRS-A, including *Pyrococcus horikoshii* (Fukunaga et al. 2005). ValRS, IleRS, and LeuRS share a post-transfer [editing domain](/superfamily/class1/Editing_domain_1a), absent from MetRS. This editing domain typically resides within the zinc finger, providing further amino acid selectivity by expelling a wide range of mistargetted amino acids such alanine, cysteine, threonine, valine, isoleucine, methionine, homocysteine, and norvaline (Gomez and Ibba, 2020). However, unlike LeuRS-A, ValRS, and IleRS, the editing domain of LeuRS-B resides downstream of the zinc finger and CP2 (Fukunaga et al. 2005). Thus, the domain appears to have hopped between the two positions, or may have originated as a trans-acting factor. LeuRS-A is lacking the connecting peptide 3 (CP3) domain found in IleRS, ValRS, and LeuRS-B. The tRNA is recognised by two C-terminal domains (Fukunaga et al. 2005). The first is a [helical domain](/superfamily/class1/Anticodon_binding_domain_CRIMVL) that recognises the anticodon binding domain, and is universal across the members of subclass Ia plus [CysRS](/class1/cys) and ArgRS. The second domain (the C-terminal domain) is unique to LeuRS-A and appears to be beneficial, but not essential, for aminoacylation activity (Fukunaga et al. 2005). LeuRS-A has been widely used for studying ancestral AARS models, such as urzymes. Urzyme constructs - which lack the editing domain, zinc finger, CP2, and C-terminal domains - still retain aminoacylation activity, albeit with much lower activity and specificity (Hobson et al. 2022, Carter and Wills 2022). Mutational studies have revealed that mutating the histidine residues of the HIGH motif of the LeuRS-A urzyme improved the enzyme's catalytic activity, suggesting that these insertions may have predated the motifs (Tang et al. 2023).

References

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