title: Selective metabolism of hypothiocyanous acid by mammalian thioredoxin reductase promotes lung innate immunity and antioxidant defense.
abstract: The endogenously produced oxidant hypothiocyanous acid (HOSCN) inhibits and kills pathogens but paradoxically is well tolerated by mammalian host tissue. Mammalian high molecular weight thioredoxin reductase (H-TrxR) is evolutionarily divergent from bacterial low molecular weight thioredoxin reductase (L-TrxR). Notably, mammalian H-TrxR contains a selenocysteine (Sec) and has wider substrate reactivity than L-TrxR. Recombinant rat cytosolic H-TrxR1, mouse mitochondrial H-TrxR2, and a purified mixture of both from rat selectively turned over HOSCN (kcat = 357 ± 16 min(-1); Km = 31.9 ± 10.3 μM) but were inactive against the related oxidant hypochlorous acid. Replacing Sec with Cys or deleting the final eight C-terminal peptides decreased affinity and turnover of HOSCN by H-TrxR. Similarly, glutathione reductase (an H-TrxR homologue lacking Sec) was less effective at HOSCN turnover. In contrast to H-TrxR and glutathione reductase, recombinant Escherichia coli L-TrxR was potently inhibited by HOSCN (IC50 = 2.75 μM). Similarly, human bronchial epithelial cell (16HBE) lysates metabolized HOSCN, but E. coli and Pseudomonas aeruginosa lysates had little or no activity. HOSCN selectively produced toxicity in bacteria, whereas hypochlorous acid was nonselectively toxic to both bacteria and 16HBE. Treatment with the H-TrxR inhibitor auranofin inhibited HOSCN metabolism in 16HBE lysates and significantly increased HOSCN-mediated cytotoxicity. These findings demonstrate both the metabolism of HOSCN by mammalian H-TrxR resulting in resistance to HOSCN in mammalian cells and the potent inhibition of bacterial L-TrxR resulting in cytotoxicity in bacteria. These data support a novel selective mechanism of host defense in mammals wherein HOSCN formation simultaneously inhibits pathogens while sparing host tissue.
authors: Department of Pharmaceutical Sciences, University of Colorado Denver, Aurora, Colorado 80045, USA.
authors: Joshua D
authors: JD
authors: Chandler
authors: David P
authors: DP
authors: Nichols
authors: Jerry A
authors: JA
authors: Nick
authors: Robert J
authors: RJ
authors: Hondal
authors: Brian J
authors: BJ
authors: Day
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D000975:Antioxidants / Q000378:metabolism*; D001310:Auranofin / Q000494:pharmacology; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D055745:Bronchioles / Q000166:cytology; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D006997:Hypochlorous Acid / Q000494:pharmacology; D007113:Immunity, Innate; D007700:Kinetics; D008168:Lung / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D050296:Microbial Viability / Q000187:drug effects; D051381:Rats; D017279:Selenocysteine / Q000235:genetics / Q000378:metabolism; D013379:Substrate Specificity; D013861:Thiocyanates / Q000378:metabolism* / Q000494:pharmacology; D013880:Thioredoxin-Disulfide Reductase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
chemical_list: D000975:Antioxidants; D001426:Bacterial Proteins; D013861:Thiocyanates; C517667:hypothiocyanous acid; D017279:Selenocysteine; D001310:Auranofin; D006997:Hypochlorous Acid; D013880:Thioredoxin-Disulfide Reductase; D003545:Cysteine
keywords: Cystic fibrosis; Infectious diseases; Inflammation; Innate immunity; Thioredoxin reductase; hypothiocyanite; thiocyanate
references: Annu Rev Biochem. 1996;65:83-100
references: 8811175
references: J Biol Chem. 1999 Sep 3;274(36):25933-44
references: 10464338
references: PLoS One. 2011;6(11):e27641
references: 22132121
references: Free Radic Biol Med. 2011 May 1;50(9):1144-50
references: 21334431
references: Biochemistry. 2006 Oct 17;45(41):12610-6
references: 17029415
references: Free Radic Biol Med. 2007 Mar 15;42(6):872-81
references: 17320769
references: J Biol Chem. 2009 Jan 9;284(2):723-7
references: 18757362
references: Biochem J. 1966 Aug;100(2):373-81
references: 4290983
references: Proc Natl Acad Sci U S A. 2009 Dec 1;106(48):20515-9
references: 19918082
references: Antiviral Res. 1995 Mar;26(2):161-71
references: 7605114
references: Biochem J. 1997 Oct 15;327 ( Pt 2):487-92
references: 9359420
references: Biochem Mol Med. 1996 Apr;57(2):134-8
references: 8733891
references: Trends Parasitol. 2002 Jul;18(7):302-8
references: 12379950
references: Eur J Biochem. 2000 Oct;267(20):6102-9
references: 11012661
references: Antioxid Redox Signal. 2000 Winter;2(4):811-20
references: 11213485
references: Anal Biochem. 2003 Jan 15;312(2):224-7
references: 12531209
references: J Leukoc Biol. 2005 May;77(5):598-625
references: 15689384
references: Respir Res. 2010 Aug 27;11:119
references: 20799947
references: J Immunol. 2006 Dec 15;177(12):8714-22
references: 17142773
references: Arch Biochem Biophys. 2006 Jan 15;445(2):235-44
references: 16125131
references: Amino Acids. 2011 Jun;41(1):73-89
references: 20397034
references: FASEB J. 2013 Apr;27(4):1394-403
references: 23248236
references: Folia Histochem Cytobiol. 2008;46(2):245-6
references: 18519245
references: Biochem Pharmacol. 2012 Dec 1;84(11):1381-7
references: 22968041
references: Biochemistry. 2006 Apr 25;45(16):5158-70
references: 16618105
references: Infect Immun. 1976 Mar;13(3):800-7
references: 5374
references: J Physiol. 2004 Nov 15;561(Pt 1):183-94
references: 15345749
references: Chem Res Toxicol. 2009 Nov;22(11):1833-40
references: 19821602
references: Nat Med. 2012 Sep;18(9):1386-93
references: 22922410
references: J Biol Chem. 2001 Jan 5;276(1):215-24
references: 11013238
references: Oral Microbiol Immunol. 1992 Oct;7(5):315-20
references: 1494457
references: PLoS One. 2011;6(9):e23637
references: 21909403
references: J Biol Chem. 2002 Mar 22;277(12):9757-62
references: 11733505
references: Infect Immun. 2008 Oct;76(10):4455-62
references: 18644874
references: J Biol Chem. 2009 Oct 16;284(42):28977-85
references: 19710012
references: Antioxid Redox Signal. 2012 Aug 1;17(3):411-21
references: 22229717
references: Antimicrob Agents Chemother. 1985 Jan;27(1):96-101
references: 2984983
references: Biochem J. 2012 Jan 1;441(1):305-16
references: 21892922
references: Methods Mol Med. 2002;70:409-31
references: 11917540
references: Biochem J. 1995 Jan 1;305 ( Pt 1):59-64
references: 7826354
references: Am J Respir Crit Care Med. 2002 May 15;165(10):1364-70
references: 12016097
references: J Dent Res. 1982 Aug;61(8):982-5
references: 6955343
references: Am J Respir Cell Mol Biol. 2003 Aug;29(2):206-12
references: 12626341
references: Biochemistry. 2010 Mar 9;49(9):2068-74
references: 20085320
references: Clin Chem. 1976 Feb;22(2):151-60
references: 2389
references: Rev Esp Enferm Dig. 2004 Nov;96(11):773-83
references: 15584851