Regulation of thioredoxin peroxidase activity by C-terminal truncation.
Thioredoxin peroxidase is a member of peroxiredoxin (Prx) family, which uses a thioredoxin (Trx) as an immediate electron donor for the reduction of peroxide. We have identified C-terminal truncated TPx from Schizosaccharomyces pombe and also have found the truncated form is significantly tenacious against the inactivation of H2O2 than the intact form. Peroxidase assay of a series of recombinant C-terminal truncation mutants (Delta192, Delta191, Delta188, Delta184, Delta176, and Delta165) revealed that TPx could be inactivated (Delta192), reactivated (Delta191-Delta176) and reinactivated (Delta165) by serial truncation from C-terminus. We did not find any significant kinetic difference among reactivated forms; however, distinctive loss of affinity to H2O2 (K(m) = 5 microM) than that of the intact form (<<5 microM, undeterminable) was monitored. Characterization of a series of Lys(191) point mutants manifested that the loss of affinity caused by a deprivation of positive charge born in Lys(191) and the loss of affinity resulted in the resistibility to H2O2. Disk inhibition assay with S. pombe cells overexpressing wild-type, Delta192 and Delta191 mutants evidenced that the truncated forms functioning in vitro as well as in vivo.
Koo
Kyung Hee
KH
Department of Biological Sciences, Chonnam National University, Gwangju, 500-757, Korea.
Lee
Songmi
S
Jeong
Soo Young
SY
Kim
Eui Tae
ET
Kim
Hyung Jung
HJ
Kim
Kanghwa
K
Song
Kiwon
K
Chae
Ho Zoon
HZ
D000595:Amino Acid Sequence; D000975:Antioxidants / Q000378:metabolism; D020134:Catalytic Domain; D003545:Cysteine / Q000378:metabolism; D004789:Enzyme Activation; D015971:Gene Expression Regulation, Enzymologic; D015966:Gene Expression Regulation, Fungal; D006861:Hydrogen Peroxide / Q000378:metabolism; D008239:Lysine / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D009363:Neoplasm Proteins; D010446:Peptide Fragments / Q000378:metabolism; D010544:Peroxidases / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D054464:Peroxiredoxins; D012568:Schizosaccharomyces / Q000201:enzymology*; D017384:Sequence Deletion
D000975:Antioxidants; D009363:Neoplasm Proteins; D010446:Peptide Fragments; D006861:Hydrogen Peroxide; D010544:Peroxidases; D054464:Peroxiredoxins; D008239:Lysine; D003545:Cysteine