Crystallization and preliminary X-ray analysis of a decameric form of cytosolic thioredoxin peroxidase 1 (Tsa1), C47S mutant, from Saccharomyces cerevisiae.
Saccharomyces cerevisiae cytosolic thioredoxin peroxidase 1 (cTPxI or Tsa1) is a bifunctional enzyme with protective roles in cellular defence against oxidative and thermal stress that exhibits both peroxidase and chaperone activities. Protein overoxidation and/or high temperatures induce great changes in its quaternary structure and lead to its assembly into large complexes that possess chaperone activity. A recombinant mutant of Tsa1 from S. cerevisiae, with Cys47 substituted by serine, was overexpressed in Escherichia coli as a His(6)-tagged fusion protein and purified by nickel-affinity chromatography. Crystals were obtained from protein previously treated with 1,4-dithiothreitol by the hanging-drop vapour-diffusion method using PEG 3000 as precipitant and sodium fluoride as an additive. Diffraction data were collected to 2.8 A resolution using a synchrotron-radiation source. The crystal structure was solved by molecular-replacement methods and structure refinement is currently in progress.
Departamento de Genética e Biologia Evolutiva, Instituto de Biociências, Universidade de São Paulo, 05508-900 São Paulo-SP, Brazil.
Marcos Antonio
MA
de Oliveira
Victor
V
Genu
Karen Fulan
KF
Discola
Simone Vidigal
SV
Alves
Luis Eduardo Soares
LE
Netto
Beatriz Gomes
BG
Guimarães
D000595:Amino Acid Sequence; D003460:Crystallization; D018360:Crystallography, X-Ray; D003545:Cysteine / Q000235:genetics; D003600:Cytosol / Q000201:enzymology*; D008969:Molecular Sequence Data; D009154:Mutation; D010544:Peroxidases / Q000737:chemistry* / Q000235:genetics; D054464:Peroxiredoxins; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics; D012694:Serine / Q000235:genetics
D029701:Saccharomyces cerevisiae Proteins; D012694:Serine; D010544:Peroxidases; D054464:Peroxiredoxins; D003545:Cysteine