Amino-acid-dependent shift in tRNA synthetase editing mechanisms

Area of Science:

• Molecular Biology
• Biochemistry
• Genetics

Background:

• Aminoacyl-tRNA synthetases (aaRS) are crucial for protein synthesis, preventing mistranslation through proofreading.
• Leucyl-tRNA synthetase (LeuRS) possesses both pre- and post-transfer editing activities, with their dominance varying.
• Yeast cytoplasmic LeuRS (ycLeuRS) can misactivate structurally similar amino acids like isoleucine and methionine.

Purpose of the Study:

• To investigate the editing mechanisms employed by yeast cytoplasmic LeuRS (ycLeuRS) when mischarging occurs.
• To determine whether pre- or post-transfer editing predominates for different noncognate amino acids.
• To elucidate the role of ycLeuRS editing in preventing mistranslation.

Main Methods:

• Enzymatic assays to measure aminoacylation and hydrolysis rates.
• Characterization of pre- and post-transfer editing activities of ycLeuRS in vitro.
• Comparative analysis of editing efficiency for isoleucine and methionine mischarging.

Main Results:

• ycLeuRS exhibits robust post-transfer editing activity for isoleucine mischarged onto tRNA(Leu).
• The post-transfer editing of methionine mischarged onto tRNA(Leu) by ycLeuRS is significantly weaker.
• Pre-transfer editing efficiently clears methionyl-adenylate, indicating a preference for this pathway when methionine is involved.

Conclusions:

• ycLeuRS utilizes both pre- and post-transfer editing pathways to maintain translational fidelity.
• The specific noncognate amino acid dictates which editing pathway predominates in ycLeuRS.
• This adaptable editing strategy ensures accurate protein synthesis by preventing mistranslation.