Allosteric networks regulate function in multi‐enzyme complexes

Abstract

Networks of noncovalent amino acid interactions propagate allosteric signals throughout proteins. Tryptophan synthase (TS) is an allosterically controlled bienzyme in which the indole product of the alpha subunit (αTS) is transferred through a 25 Å hydrophobic tunnel to the active site of the beta subunit (βTS). Previous nuclear magnetic resonance and molecular dynamics simulations identified allosteric networks in αTS important for its function. Here, we show that these allosteric networks change across the catalytic cycle of αTS, such that chemical catalysis is associated with specific conformational dynamics and networks not observed in the substrate‐ and products‐bound states. Moreover, substitution of a distant, surface‐exposed network residue in αTS enhances tryptophan production, not by activating αTS function, but through dynamically controlling the opening of the indole channel and stimulating βTS activity. While stimulation is modest, the substitution also enhances cell growth in a tryptophan‐auxotrophic strain of Escherichia coli compared to complementation with wild‐type αTS, emphasizing the biological importance of the network. Network residues thus represent ideal targets to optimize mutagenesis strategies in protein engineering studies directed towards changing enzyme function and regulation, especially in multi‐enzyme complexes.