Crowder architecture matters: Distinct effects of linear and branched macromolecular crowders on dynamics and function of Escherichia coli prolyl-tRNA synthetase

Abstract

In vitro enzyme studies often neglect the highly crowded nature of the cellular interior. Using spectroscopy and molecular dynamics simulations we investigated how crowders of variable architecture, namely, linear polyethylene glycols and branched polysaccharides (dextran 40 and ficoll 70) modulate the conformational landscape and function of the modular Escherichia coli prolyl tRNA synthetase. This work reveals that all crowders favor a compact enzyme conformation but modulate conformational dynamics and substrate binding through distinct mechanisms, providing insights relevant to protein stabilization strategies and the rational design of therapeutics targeting modular enzymes.