Bioluminescent detection of peroxynitrite with a boronic acid-caged luciferin. Free Radic Biol Med 2013 Aug;61:40-50
Date
03/12/2013Pubmed ID
23474271Pubmed Central ID
PMC3795912DOI
10.1016/j.freeradbiomed.2013.02.020Scopus ID
2-s2.0-84878846249 (requires institutional sign-in at Scopus site) 40 CitationsAbstract
Peroxynitrite, a highly reactive biological oxidant, is formed under pathophysiologic conditions from the diffusion-limited reaction of nitric oxide and superoxide radical anion. Peroxynitrite has been implicated as the mediator of nitric oxide toxicity in many diseases and as an important signaling disrupting molecule (L. Liaudet et al., Front. Biosci.14, 4809-4814, 2009) [1]. Biosensors effective at capturing peroxynitrite in a specific and fast enough manner for detection, along with readouts compatible with in vivo studies, are lacking. Here we report that the boronic acid-based bioluminescent system PCL-1 (peroxy-caged luciferin-1), previously reported as a chemoselective sensor for hydrogen peroxide (G.C. Van de Bittner et al., Proc. Natl. Acad. Sci. USA107, 21316-21321, 2010) [2], reacts with peroxynitrite stoichiometrically with a rate constant of 9.8±0.3×10(5)M(-1)s(-1) and a bioluminescence detection limit of 16nM, compared to values of 1.2±0.3M(-1)s(-1) and 231nM for hydrogen peroxide. Further, we demonstrate bioluminescent detection of peroxynitrite in the presence of physiological competitors: carbon dioxide, glutathione, albumin, and catalase. We also demonstrate the utility of this method to assess peroxynitrite formation in mammalian cells by measuring peroxynitrite generated under normal culture conditions after stimulation of macrophages with bacterial endotoxin lipopolysaccharide. Thus, the PCL-1 method for measuring peroxynitrite generation shows superior selectivity over other oxidants under in vivo conditions.
Author List
Sieracki NA, Gantner BN, Mao M, Horner JH, Ye RD, Malik AB, Newcomb ME, Bonini MGAuthor
Benjamin N. Gantner PhD Assistant Professor in the Medicine department at Medical College of WisconsinMESH terms used to index this publication - Major topics in bold
AnimalsBenzothiazoles
Boronic Acids
Luminescent Measurements
Mice
Nitric Oxide Synthase Type II
Peroxynitrous Acid
