Crystallization and preliminary X-ray diffraction analysis of NADPH-dependent thioredoxin reductase I from Saccharomyces cerevisiae.
Thioredoxin reductase 1 (Trr1) from Saccharomyces cerevisiae is a member of the family of pyridine nucleotide-disulfide oxidoreductases capable of reducing the redox-active disulfide bond of the cytosolic thioredoxin 1 (Trx1) and thioredoxin 2 (Trx2). NADPH, Trr1 and Trx1 (or Trx2) comprise the thioredoxin system, which is involved in several biological processes, including the reduction of disulfide bonds and response to oxidative stress. Recombinant Trr1 was expressed in Escherichia coli as a His6-tagged fusion protein and purified by nickel-affinity chromatography. The protein was crystallized using the hanging-drop vapour-diffusion method in the presence of PEG 3000 as precipitant after treatment with hydrogen peroxide. X-ray diffraction data were collected to a maximum resolution of 2.4 A using a synchrotron-radiation source. The crystal belongs to the centred monoclinic space group C2, with unit-cell parameters a = 127.97, b = 135.41, c = 75.81 A, beta = 89.95 degrees. The crystal structure was solved by molecular-replacement methods and structure refinement is in progress.
Oliveira
Marcos Antonio de
MA
Departamento de Biologia, Instituto de Biociências, Universidade de São Paulo, Rua do Matão 277, São Paulo SP, Brazil.
Discola
Karen Fulan
KF
Alves
Simone Vidigal
SV
Barbosa
João Alexandre Ribeiro Gonçalves
JA
Medrano
Francisco Javier
FJ
Netto
Luis Eduardo Soares
LE
Guimarães
Beatriz Gomes
BG
D003001:Cloning, Molecular; D003460:Crystallization / Q000379:methods; D006639:Histidine; D011092:Polyethylene Glycols; D016133:Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry*; D013880:Thioredoxin-Disulfide Reductase / Q000737:chemistry*; D014835:Volatilization; D014961:X-Ray Diffraction
D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D011092:Polyethylene Glycols; D006639:Histidine; D013880:Thioredoxin-Disulfide Reductase