Thioredoxin-2 but not thioredoxin-1 is a substrate of thioredoxin peroxidase-1 from Drosophila melanogaster -: Isolation and characterization of a second thioredoxin in D-melanogaster and evidence for distinct biological functions of Trx-1 and Trx-2

被引:73
作者
Bauer, H [1 ]
Kanzok, SM [1 ]
Schirmer, RH [1 ]
机构
[1] Univ Heidelberg, Biochem Ctr, D-69120 Heidelberg, Germany
关键词
D O I
10.1074/jbc.M200636200
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
As Drosophila melanogaster does not contain glutathione reductase, the thioredoxin system has a key function for glutathione disulfide reduction in insects (Kanzok, S. M., Fechner, A., Bauer, H., Ulschmid, J. K., Muller, H. M., Botella-Munoz, J., Schneuwly, S., Schirmer, R. H., and Becker, K. (2001) Science 291, 643-646). In view of these unique conditions, the protein systems participating in peroxide metabolism and in redox signaling are of special interest. The genes for a second thioredoxin (DmTrx-2) and a thioredoxin peroxidase (DmTPx-1) were cloned and expressed, and the proteins were characterized. In its disulfide form, the 13-kDa protein thioredoxin-2 is a substrate of thioredoxin reductase-1 (K-m=5.2 muM, k(cat)=14.5 s(-1)) and in its dithiol form, an electron donor for TPx-1 (K-m=9 muM, k(cat)=5.4 s(-1)). DmTrx-2 is capable of reducing glutathione disulfide with a second order rate constant of 170 M-1 s(-1) at pH 7.4 and 25degreesC. Western blot analysis indicated that this thioredoxin represents up to 1% of the extractable protein of D. melanogaster Schneider cells or whole fruit flies. Recombinant thioredoxin peroxidase-1 (subunit molecular mass=23 kDa) was found to be a decameric protein that can efficiently use Trx-2 but not Trx-1 as a reducing substrate. The new electron pathway found in D. melanogaster is also representative for insects that serve as vectors of disease. As a first step we have cloned and functionally expressed the gene that is the orthologue of DmTrx-2 in the malaria mosquito Anopheles gambiae.
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页码:17457 / 17463
页数:7
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