A live bacteria enzyme assay for identification of human disease mutations and drug screening

Abstract

Advances in high-throughput sequencing have enabled the identification of genetic variations associated with human disease. However, deciphering the functional significance of these variations remains challenging. Here we propose an alternative approach that uses humanized Escherichia coli to study human genetic enzymopathies and to screen candidate drug effects on metabolic targets. By replacing selected E. coli metabolic enzymes with their human orthologues and their sequence variants, we demonstrate that the growth rate of E. coli reflects the in vivo activity of heterologously expressed human enzymes. This approach accurately reflected enzyme activities of known sequence variants, enabling rapid screening of causal sequence variations associated with human diseases. This approach bridges the gap between in vitro assays and cell-based assays. Our findings suggest that the proposed approach using a humanized E. coli strain holds promise for drug discovery, offering a high-throughput and cost-effective platform for identifying new compounds targeting human enzymes. Continued research and innovation in this field have the potential to impact the development and practice of precision medicine.