posted on 2020-03-19, 18:39authored byJacek Kozuch, Samuel H. Schneider, Steven G. Boxer
A biophysical
understanding of the mechanistic, chemical, and physical
origins underlying antibiotic action and resistance is vital to the
discovery of novel therapeutics and the development of strategies
to combat the growing emergence of antibiotic resistance. The site-specific
introduction of stable-isotope labels into chemically complex natural
products is particularly important for techniques such as NMR, IR,
mass spectrometry, imaging, and kinetic isotope effects. Toward this
goal, we developed a biosynthetic strategy for the site-specific incorporation
of 13C labels into the canonical β-lactam carbonyl
of penicillin G and cefotaxime, the latter via cephalosporin C. This
was achieved through sulfur-replacement with 1-13C-l-cysteine, resulting in high isotope incorporations and milligram-scale
yields. Using 13C NMR and isotope-edited IR difference
spectroscopy, we illustrate how these molecules can be used to interrogate
interactions with their protein targets, e.g., TEM-1 β-lactamase.
This method provides a feasible route to isotopically labeled penicillin
and cephalosporin precursors for future biophysical studies.