# Expanding the Genetic Code with Lysine Aminoacylation

## TL;DR

This study expands the genetic code to create tools for studying lysine aminoacylation and shows how it affects metabolic enzymes in human cells.

## Contribution

The development of orthogonal translation systems for ten types of lysine aminoacylation in bacterial and mammalian cells.

## Key findings

- Lysine valylation and tyrosylation impair pyruvate kinase and glucose-6-phosphate dehydrogenase activities.
- Lysine valylation of pyruvate kinase decreases the basal glycolytic rate in living human cells.

## Abstract

Lysine aminoacylation
is a newly discovered protein post-translational
modification that is found in humans. However, few studies have been
implemented to further investigate its function, possibly due to limited
tools to produce target proteins with homogeneously aminoacylated
lysine residues at specific sites. To achieve this goal, we applied
the genetic code expansion strategy, engineered pyrrolysyl-tRNA synthetase,
and established orthogonal translation systems for ten types of lysine
aminoacylation compatible for both bacterial and mammalian cells.
Because metabolic enzymes are preferred substrate proteins of lysine
aminoacylation, we tested the effect of lysine aminoacylation on metabolic
enzymes and demonstrated that lysine valylation and tyrosylation impaired
pyruvate kinase and glucose-6-phosphate dehydrogenase activities,
respectively. Further in vivo studies showed that
lysine valylation of pyruvate kinase decreased the basal glycolytic
rate in living human cells. In summary, this work provides a toolbox
to study lysine aminoacylation.