Immuno-spin trapping of a post-translational carboxypeptidase B1 radical formed by a dual role of xanthine oxidase and endothelial nitric oxide synthase in acute septic mice

Post-translational modification of proteins due to exposure to radicals and other reactive species are markers of metabolic and inflammatory oxidative stress Such as sepsis. This study uses the nitrone spin-trap DMPO and a combination of immuno-spin trapping and mass spectrometry to identify in vivo products of radical reactions in mice. We report the detection of dose-dependent production of DMPO-carboxypeptidase B1 (CPB1) adducts in the spleens of mice treated with lipopolysaccharide (LPS). Additionally, we report significant detection of DMPO-CPBI adducts in mice experiencing normal physiological conditions. Treatments with inhibitors and experiments with knock-out Mice indicate that xanthine oxidase and endothelial nitric oxide synthase are important sources of the reactive species that lead to CPBI adduct formation. We also report a significant loss of CPB1 activity following LPS challenge in conjunction with an increase in CPBI protein accumulation. This Suggests the presence of a possible mechanism for CPBI activity loss with compensatory protein production. Published by Elsevier Inc.