Connecting glutamine synthetase biophysics to the cellular environment

Enzymes permit life to exist outside of thermodynamic equilibrium and play a crucial role in numerous diseases; however, our understanding of how these enzymes function in vivo is hampered by limitations applying in vitro measurements to the intracellular environment. Here, we present groundwork to establish an in vivo metric of enzyme specific activity as applied to a model enzyme, human glutamine synthetase (GS). Utilizing select clinically relevant variants, in vitro kinetic and cryo-EM structural analysis of GS has permitted a foundational understanding of GS catalytic mechanism from which to ground in vivo analyses.