Inter-domain redox communication in flavoenzymes of the quiescin/sulfhydryl oxidase family: Role of a thioredoxin domain in disulfide bond formation

被引:63
作者
Raje, S [1 ]
Thorpe, C [1 ]
机构
[1] Univ Delaware, Dept Biochem & Chem, Newark, DE 19716 USA
关键词
D O I
10.1021/bi030003z
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Flavoproteins of the quiescin/sulfhydryl oxidase (QSOX) family catalyze oxidation of peptide and protein thiols to disulfides with the reduction of oxygen to hydrogen peroxide. QSOX family members contain several domains, including an N-terminal thioredoxin domain (Trx) and an FAD-binding-domain (ERV) toward the C-terminus. Partial proteolysis of avian QSOX leads to two fragments, designated 30 and 60 kDa from their apparent mobilities on SDS-PAGE. The 30 kDa fragment is a monomer under nondenaturing conditions and contains a Trx domain with a CxxC sequence typical of protein disulfide isomerase (WCGHC). This QSOX fragment is not detectably glycosylated, contains no detectable FAD, and shows undetectable sulfhydryl oxidase activity. In contrast, the 60 kDa fragment is a dimeric glycoprotein that binds FAD tightly and oxidizes dithiothreitol about 1000-fold slower than intact QSOX. Reduced RNase is not a significant substrate of the 60 kDa fragment. The redox behavior of the 60 kDa flavoprotein fragment is profoundly different from that of intact QSOX. Thus, dithionite or photochemical reduction of the 60 kDa fragment leads to two-electron reduction of the FAD without subsequent reduction of the other two CxxC motifs or the appearance of a thiolate to flavin charge-transfer complex. Further characterization of the fragments and insights gained from the crystal structure of yeast ERV2p (Gross, E., Sevier, C. S., Vala, A., Kaiser, C. A., and Fass, D. (2002) Nat. Struct. Biol. 9, 61-67) suggest that the flow of reducing equivalents in intact avian QSOX is dithiol substrate --> C80/83 --> C519/522 --> C459/462 --> FAD --> oxygen. The ancient fusion of thioredoxin domains to a catalytically more limited ERV domain has produced an efficient catalyst for the direct introduction of disulfide bonds into a wide range of proteins and peptides in multicellular organisms.
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页码:4560 / 4568
页数:9
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共 37 条
  • [1] Rat seminal vesicle FAD-dependent sulfhydryl oxidase -: Biochemical characterization and molecular cloning of a member of the new sulfhydryl oxidase/quiescin Q6 gene family
    Benayoun, B
    Esnard-Fève, A
    Castella, S
    Courty, Y
    Esnard, F
    [J]. JOURNAL OF BIOLOGICAL CHEMISTRY, 2001, 276 (17) : 13830 - 13837
  • [2] The CXXC motif: A rheostat in the active site
    Chivers, PT
    Prehoda, KE
    Raines, RT
    [J]. BIOCHEMISTRY, 1997, 36 (14) : 4061 - 4066
  • [3] COPPOCK DL, 1993, CELL GROWTH DIFFER, V4, P483
  • [4] The quiescin Q6 gene (QSCN6) is a fusion of two ancient gene families: Thioredoxin and ERV1
    Coppock, DL
    Cina-Poppe, D
    Gilleran, S
    [J]. GENOMICS, 1998, 54 (03) : 460 - 468
  • [5] Gailer J, 1999, APPL ORGANOMET CHEM, V13, P837, DOI 10.1002/(SICI)1099-0739(199911)13:11<837::AID-AOC924>3.0.CO
  • [6] 2-Z
  • [7] Yeast Erv2p is the first microsomal FAD-linked sulfhydryl oxidase of the Erv1p/Alrp protein family
    Gerber, J
    Mühlenhoff, U
    Hofhaus, G
    Lill, R
    Lisowsky, T
    [J]. JOURNAL OF BIOLOGICAL CHEMISTRY, 2001, 276 (26) : 23486 - 23491
  • [8] A new FAD-binding fold and intersubunit disulfide shuttle in the thiol oxidase Erv2p
    Gross, E
    Sevier, CS
    Vala, A
    Kaiser, CA
    Fass, D
    [J]. NATURE STRUCTURAL BIOLOGY, 2002, 9 (01) : 61 - 67
  • [9] Egg white sulfhydryl oxidase: Kinetic mechanism of the catalysis of disulfide bond formation
    Hoober, KL
    Thorpe, C
    [J]. BIOCHEMISTRY, 1999, 38 (10) : 3211 - 3217
  • [10] Homology between egg white sulfhydryl oxidase and quiescin Q6 defines a new class of flavin-linked sulfhydryl oxidases
    Hoober, KL
    Glynn, NM
    Burnside, J
    Coppock, DL
    Thorpe, C
    [J]. JOURNAL OF BIOLOGICAL CHEMISTRY, 1999, 274 (45) : 31759 - 31762