The genotype‐phenotype landscape of an allosteric protein

Allostery is a fundamental biophysical mechanism that underlies cellular sensing, signaling, and metabolism. Yet a quantitative understanding of allosteric genotype‐phenotype relationships remains elusive. Here, we report the large‐scale measurement of the genotype‐phenotype landscape for an allosteric protein: the lac repressor from Escherichia coli, LacI. Using a method that combines long‐read and short‐read DNA sequencing, we quantitatively measure the dose‐response curves for nearly 10(5) variants of the LacI genetic sensor. The resulting data provide a quantitative map of the effect of amino acid substitutions on LacI allostery and reveal systematic sequence‐structure‐function relationships. We find that in many cases, allosteric phenotypes can be quantitatively predicted with additive or neural‐network models, but unpredictable changes also occur. For example, we were surprised to discover a new band‐stop phenotype that challenges conventional models of allostery and that emerges from combinations of nearly silent amino acid substitutions.