Inactivation of human salivary glutathione transferase P1-1 by hypothiocyanite: a post-translational control system in search of a role.
Glutathione transferases (GSTs) are a superfamily of detoxifying enzymes over-expressed in tumor tissues and tentatively proposed as biomarkers for localizing and monitoring injury of specific tissues. Only scarce and contradictory reports exist about the presence and the level of these enzymes in human saliva. This study shows that GSTP1-1 is the most abundant salivary GST isoenzyme, mainly coming from salivary glands. Surprisingly, its activity is completely obscured by the presence of a strong oxidizing agent in saliva that causes a fast and complete, but reversible, inactivation. Although salivary α-defensins are also able to inhibit the enzyme causing a peculiar half-site inactivation, a number of approaches (mass spectrometry, site directed mutagenesis, chromatographic and spectrophotometric data) indicated that hypothiocyanite is the main salivary inhibitor of GSTP1-1. Cys47 and Cys101, the most reactive sulfhydryls of GSTP1-1, are mainly involved in a redox interaction which leads to the formation of an intra-chain disulfide bridge. A reactivation procedure has been optimized and used to quantify GSTP1-1 in saliva of 30 healthy subjects with results of 42±4 mU/mg-protein. The present study represents a first indication that salivary GSTP1-1 may have a different and hitherto unknown function. In addition it fulfills the basis for future investigations finalized to check the salivary GSTP1-1 as a diagnostic biomarker for diseases.
Department of Chemical Sciences and Technologies, University of Rome "Tor Vergata", 00133 Rome, Italy.
Raffaele
R
Fabrini
Department of Chemical Sciences and Technologies, University of Rome "Tor Vergata", 00133 Rome, Italy.
Alessio
A
Bocedi
Department of Cell Biology and Neurosciences, Istituto Superiore di Sanità, 00161 Rome, Italy.
Serena
S
Camerini
Department of Applied Clinical Sciences, University of L'Aquila, 67100 L'Aquila, Italy.
Marco
M
Fusetti
Department of Clinical Sciences and Translational Medicine, University of Rome "Tor Vergata", 00133 Rome, Italy.
Fabrizio
F
Ottaviani
Department of Clinical Sciences and Translational Medicine, University of Rome "Tor Vergata", 00133 Rome, Italy.
Francesco M
FM
Passali
Department of Clinical Sciences and Translational Medicine, University of Rome "Tor Vergata", 00133 Rome, Italy.
Davide
D
Topazio
Institute of Biochemistry and Clinical Biochemistry, Catholic University and/or Institute for Molecular Recognition, National Research Council, 00168 Rome, Italy.
Federica
F
Iavarone
Department of Chemical Sciences and Technologies, University of Rome "Tor Vergata", 00133 Rome, Italy.
Irene
I
Francia
Institute of Biochemistry and Clinical Biochemistry, Catholic University and/or Institute for Molecular Recognition, National Research Council, 00168 Rome, Italy.
Massimo
M
Castagnola
Department of Chemical Sciences and Technologies, University of Rome "Tor Vergata", 00133 Rome, Italy.
Giorgio
G
Ricci
D000328:Adult; D000368:Aged; D000890:Anti-Infective Agents / Q000494:pharmacology; D015415:Biomarkers / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005260:Female; D051549:Glutathione S-Transferase pi / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006801:Humans; D008297:Male; D008875:Middle Aged; D012463:Saliva / Q000201:enzymology*; D012471:Salivary Proteins and Peptides / Q000037:antagonists & inhibitors* / Q000378:metabolism; D013861:Thiocyanates / Q000494:pharmacology*
D000890:Anti-Infective Agents; D015415:Biomarkers; D004791:Enzyme Inhibitors; D012471:Salivary Proteins and Peptides; D013861:Thiocyanates; C014607:hypothiocyanite ion; C496556:GSTP1 protein, human; D051549:Glutathione S-Transferase pi