Chemical Bioconjugation of Proteins in an Undergraduate Lab: One-Pot Oxidation and Derivatization of the N-Terminus

A laboratory experiment introducing the concept of chemical bioconjugation of proteins to undergraduate students in a therapeutically relevant context was developed. Initially, students installed an aldehyde functionality into a protein via the oxidation of the N-terminal threonine residue of the cholera toxin subunit B (CTB) protein, which was followed by subsequent modification via hydrazone addition under mild conditions with a chromophore bearing a distinct UV–vis-absorption peak. Students determined the yield of the reaction to be ca. 11% by HPLC coupled to UV–vis spectroscopy and developed key skills such as preparation of stock solutions, chemical manipulation of proteins, and analysis via HPLC. The reported experiment can be readily adapted for use with other proteins and may contribute to enhancing constructive alignment in interdisciplinary degree programs at the chemistry–biology interface.