Genetic Code Expansion in Probiotics Enables the Secretion of Covalent Protein Drugs in Mice

Oral delivery of therapeutic proteins remains a formidable
challenge.
Although engineered microbes have emerged as promising platforms for
localized drug synthesis in the gut, their functional capacity has
been restricted to the 20 canonical amino acids, limiting the chemical
diversity of biologic payloads. Here, we demonstrate that integrating
genetic code expansion (GCE) into bacterial therapy overcomes this
fundamental constraint. We engineered the probiotic Escherichia coli Nissle 1917 (EcN) to incorporate
the noncanonical amino acid fluorosulfate-l-tyrosine (FSY),
enabling in situ secretion of a site-specifically
modified covalent anti-IL-23 nanobody exhibiting picomolar binding
potency (5.9 pM). Oral administration of this engineered EcN strain,
followed by FSY supplementation, significantly ameliorated colitis
in a murine model. This approach thereby establishes a versatile and
generalizable platform that substantially expands the functional scope
and therapeutic potential of live biotherapeutics.