+ Site Statistics
+ Search Articles
+ PDF Full Text Service
How our service works
Request PDF Full Text
+ Follow Us
Follow on Facebook
Follow on Twitter
Follow on LinkedIn
+ Subscribe to Site Feeds
Most Shared
PDF Full Text
+ Translate
+ Recently Requested

Avian sulfhydryl oxidase is not a metalloenzyme: adventitious binding of divalent metal ions to the enzyme



Avian sulfhydryl oxidase is not a metalloenzyme: adventitious binding of divalent metal ions to the enzyme



Biochemistry 42(37): 11074-11082



Metal- and flavin-dependent sulfhydryl oxidases catalyze the generation of disulfide bonds with reduction of oxygen to hydrogen peroxide. The mammalian skin enzyme has been reported to be copper-dependent, but a recent protein sequence shows it belongs to the Quiescin/sulfhydryl oxidase (QSOX) flavoprotein family. This work demonstrates that avian QSOX is not a metalloenzyme, and that copper and zinc ions inhibit the oxidation of reduced pancreatic ribonuclease by the enzyme. Studies with Zn(2+), as a redox inactive surrogate for copper, show that one Zn(2+) binds to four-electron-reduced QSOX by diverting electrons away from the flavin and into two of the three redox active disulfide bridges in the enzyme. The resulting zinc complex is modestly air-stable, reverting to a spectrum of the native protein with a t(1/2) of 40 min, whereas the four-electron-reduced native QSOX is reoxidized in less than a second under comparable conditions. Using tris(2-carboxyethyl)phosphine hydrochloride (TCEP), an alternate substrate of QSOX that binds Zn(2+) relatively weakly (unlike dithiothreitol), allows rapid inhibition of oxidase activity to be demonstrated at low micromolar metal levels. Zinc binding was followed by rapid-scanning spectrophotometry. Copper also binds the four-electron-reduced form of QSOX with a visible spectrum suggestive of active site occupancy. In addition to interactions with the reduced enzyme, dialysis experiments show that multiple copper and zinc ions can bind to the oxidized enzyme without the perturbation of the flavin spectrum seen earlier. These data suggest that a reinvestigation of the metal content of skin sulfhydryl oxidases is warranted. The redox-modulated binding of zinc to QSOX is considered in light of evidence for a role of zinc-thiolate interactions in redox signaling and zinc mobilization.

Please choose payment method:






(PDF emailed within 0-6 h: $19.90)

Accession: 009775505

Download citation: RISBibTeXText

PMID: 12974644

DOI: 10.1021/bi0301385


Related references

Estimation of the distance between sulfhydryl groups S1 and specific binding centers for divalent metal ions in myosin. Biofizika 26(5): 920-923, 1981

Sulfhydryl content of bovine eye lens leucine aminopeptidase. Determination of the reactivity of the sulfhydryl groups of the zinc metalloenzyme, of the enzyme activated by Mg2+, Mn2+, and Co2+, and of the metal-free apoenzyme. Journal of Biological Chemistry 257(12): 7086-7091, 1982

Complexation and divalent of desferricoprogen with trivalent Fe, Al, Ga, In and divalent Fe, Ni, Cu, Zn metal ions: effects of the linking chain structure on the metal binding ability of hydroxamate based siderophores. Journal of Inorganic Biochemistry 98(11): 1957-1966, 2004

Unusually wide co-factor tolerance in a metalloenzyme; divalent metal ions modulate endo-exonuclease activity in T5 exonuclease. Nucleic Acids Research 29(13): 2772-2779, 2001

Probing the role of divalent metal ions in a bacterial psychrophilic metalloprotease: binding studies of an enzyme in the crystalline state by x-ray crystallography. Journal of Bacteriology 185(14): 4195-4203, 2003

The calmodulin-dependent activation and deactivation of the phosphoprotein phosphatase, calcineurin, and the effect of nucleotides, pyrophosphate, and divalent metal ions. Identification of calcineurin as a Zn and Fe metalloenzyme. Journal of Biological Chemistry 259(14): 8847-8856, 1984

The calmodulin dependent activation and deactivation of the phosphoprotein phosphatase calcineurin and the effect of nucleotides pyrophosphate and divalent metal ions identification of calcineurin as a zinc and iron metalloenzyme. Journal of Biological Chemistry 259(14): 8847-8856, 1984

Metal ions and phosphate binding in the H-N-H motif: crystal structures of the nuclease domain of ColE7/Im7 in complex with a phosphate ion and different divalent metal ions. Protein Science 11(12): 2947-2957, 2002

Interactions of dna with divalent metal ions 3. extent of metal binding experiment and theory. Biopolymers 21(1): 219-232, 1982

Interactions of DNA with divalent metal ions. III. Extent of metal binding: experiment and theory. Biopolymers 21(1): 219-232, 1982

Metal ions as cofactors for the binding of inhibitors to methionine aminopeptidase: a critical view of the relevance of in vitro metalloenzyme assays. Angewandte Chemie 44(23): 3620-3623, 2005

Investigation of the affinity and selectivity of avian prion hexarepeat peptides for physiological divalent metal ions. Journal of Inorganic Biochemistry 101(5): 783-788, 2007

A comparative study of dna interaction with divalent metal ions metal ion binding sites and dna conformational changes studied by laser raman spectroscopy. Trace Elements in Medicine 7(2): 102, 1990

Divalent metal ions binding properties of goat serum mannose binding lectin. International Journal of Biological Macromolecules 80: 324-327, 2015

Effects of divalent metal ions as modulators of enzyme reactions. Journal of Inorganic Biochemistry 51(1-2): 259-0, 1993