2NVK

Crystal Structure of Thioredoxin Reductase from Drosophila melanogaster

Summary for 2NVK

• Entry DOI: 10.2210/pdb2nvk/pdb
• Descriptor: Thioredoxin Reductase, FLAVIN-ADENINE DINUCLEOTIDE, NADP NICOTINAMIDE-ADENINE-DINUCLEOTIDE PHOSPHATE, ... (4 entities in total)
• Functional Keywords: oxidoreductase, rossmann, flavoprotein
• Biological source: Drosophila melanogaster (fruit fly)
• Total number of polymer chains: 1
• Total formula weight: 54521.33

Authors

Eckenroth, B.E.,Rould, M.A.,Hondal, R.J.,Everse, S.J. (deposition date: 2006-11-13, release date: 2007-04-24, Last modification date: 2024-10-30)

Primary citation

Eckenroth, B.E.,Rould, M.A.,Hondal, R.J.,Everse, S.J.
Structural and Biochemical Studies Reveal Differences in the Catalytic Mechanisms of Mammalian and Drosophila melanogaster Thioredoxin Reductases.
Biochemistry, 46:4694-4705, 2007

PubMed Abstract: Thioredoxin reductase (TR) from Drosophila melanogaster (DmTR) is a member of the glutathione reductase (GR) family of pyridine nucleotide disulfide oxidoreductases and catalyzes the reduction of the redox-active disulfide bond of thioredoxin. DmTR is notable for having high catalytic activity without the presence of a selenocysteine (Sec) residue (which is essential for the mammalian thioredoxin reductases). We report here the X-ray crystal structure of DmTR at 2.4 A resolution (Rwork = 19.8%, Rfree = 24.7%) in which the enzyme was truncated to remove the C-terminal tripeptide sequence Cys-Cys-Ser. We also demonstrate that tetrapeptides equivalent to the oxidized C-terminal active sites of both mouse mitochondrial TR (mTR3) and DmTR are substrates for the truncated forms of both enzymes. This truncated enzyme/peptide substrate system examines the kinetics of the ring-opening step that occurs during the enzymatic cycle of TR. The ring-opening step is 300-500-fold slower when Sec is replaced with Cys in mTR3 when using this system. Conversely, when Cys is replaced with Sec in DmTR, the rate of ring opening is only moderately increased (5-36-fold). Structures of these tetrapeptides were oriented in the active site of both enzymes using oxidized glutathione bound to GR as a template. DmTR has a more open tetrapeptide binding pocket than the mouse enzyme and accommodates the peptide Ser-Cys-Cys-Ser(ox) in a cis conformation that allows for the protonation of the leaving-group Cys by His464', which helps to explain why this TR can function without the need for Sec. In contrast, mTR3 shows a narrower pocket. One possible result of this narrower interface is that the mammalian redox-active tetrapeptide Gly-Cys-Sec-Gly may adopt a trans conformation for a better fit. This places the Sec residue farther away from the protonating histidine residue, but the lower pKa of Sec in comparison to that of Cys eliminates the need for Sec to be protonated.

PubMed: 17385893
DOI: 10.1021/bi602394p

Experimental method

X-RAY DIFFRACTION (2.4 Å)