ADP-ribosylating toxin enzymes break NADH (Figure1) and transfer the ADP-ribosyl group to a residue on a target protein, permanently inactivating or denaturing the protein. This activity is typically detected with a radioassay, which is expensive and requires radioactive materials.
ADP-ribosylation corresponds with the release of nicotinamide. It is possible to detect nicotinamide with a Gas Chromatograph/Mass Spectrometer (GC/MS) (Jacobson, Dame, Pyrek & Jacobson, 1995). The purpose of this study is to measure ADP-ribosylation activity using GC/MS by detecting the liberated nicotinamide. By derivatizing the nicotinamide, the detection limit was lowered to 0.5ng/μl. Control measurements of ADP-ribosylation activity by a cholera toxin protein found low levels of nicotinamide contamination.
CU Commons Citation
Lermo, Lindsay R.; Munson, Thomas O.; and Godsey, Michael H., "Development of a Novel GC/MS Method for the Detection of Nicotinamide and Activity of ADP-Ribosylating Toxins" (2011). Math & Science Department (SURI). 1.