PMID- 12974644
OWN - NLM
STAT- MEDLINE
DCOM- 20031028
LR  - 20131121
IS  - 0006-2960 (Print)
IS  - 0006-2960 (Linking)
VI  - 42
IP  - 37
DP  - 2003 Sep 23
TI  - Avian sulfhydryl oxidase is not a metalloenzyme: adventitious binding of divalent 
      metal ions to the enzyme.
PG  - 11074-82
AB  - 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.
FAU - Brohawn, Stephen G
AU  - Brohawn SG
AD  - Department of Chemistry and Biochemistry, University of Delaware, Newark, 
      Delaware 19716, USA.
FAU - Miksa, Irina Rudik
AU  - Miksa IR
FAU - Thorpe, Colin
AU  - Thorpe C
LA  - eng
GR  - GM 26643/GM/NIGMS NIH HHS/United States
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - United States
TA  - Biochemistry
JT  - Biochemistry
JID - 0370623
RN  - 0 (Metals)
RN  - 0 (Phosphines)
RN  - 22OAC2MO2S (tris(2-carboxyethyl)phosphine)
RN  - 789U1901C5 (Copper)
RN  - EC 1.- (Oxidoreductases)
RN  - EC 1.8.3.- (sulfhydryl oxidase)
RN  - EC 3.1.27.5 (Ribonuclease, Pancreatic)
RN  - J41CSQ7QDS (Zinc)
SB  - IM
MH  - Amino Acid Sequence
MH  - Animals
MH  - Binding Sites
MH  - Birds
MH  - Copper/chemistry
MH  - Electrons
MH  - Metals/chemistry
MH  - Mice
MH  - Models, Chemical
MH  - Molecular Sequence Data
MH  - Oxidation-Reduction
MH  - Oxidoreductases/*chemistry
MH  - Phosphines/chemistry
MH  - Protein Binding
MH  - Protein Structure, Tertiary
MH  - Ribonuclease, Pancreatic/metabolism
MH  - Sequence Homology, Amino Acid
MH  - Spectrophotometry
MH  - Temperature
MH  - Time Factors
MH  - Zinc/chemistry
EDAT- 2003/09/17 05:00
MHDA- 2003/10/29 05:00
CRDT- 2003/09/17 05:00
PHST- 2003/09/17 05:00 [pubmed]
PHST- 2003/10/29 05:00 [medline]
PHST- 2003/09/17 05:00 [entrez]
AID - 10.1021/bi0301385 [doi]
PST - ppublish
SO  - Biochemistry. 2003 Sep 23;42(37):11074-82. doi: 10.1021/bi0301385.