Purification, molecular cloning, and characterization of TRP32, a novel thioredoxin-related mammalian protein of 32 kDa.
We purified a protein of 32 kDa from human thymoma HPB-ALL cells that was co-purified with a catalytic fragment of MST (mammalian STE-20-like), a kinase of the STE20 family, which is proteolytically activated by caspase in apoptosis (Lee, K.-K., Murakawa, M., Nishida, E., Tsubuki, S., Kawashima, S., Sakamaki, K., and Yonehara, S. (1998) Oncogene 16, in press). Molecular cloning of the gene encoding this 32-kDa protein (TRP32) reveals that it is a novel protein of 289 amino acid residues and contains an NH2-terminal thioredoxin domain with a conserved thioredoxin active site. The human and mouse TRP32 proteins have 99% homology, and the thioredoxin domains are completely identical. The thioredoxin domain of TRP32 has thioredoxin-like reducing activity, which can reduce the interchain disulfide bridges of insulin in vitro. However, the thioredoxin domain of TRP32 is more sensitive to oxidation than human thioredoxin. Northern blot analysis showed that TRP32 is expressed in all human tissues. Expression of TRP32 was also confirmed in all mammalian cell lines tested by Western blot analysis using anti-TRP32 monoclonal antibody. Subcellular fractionation and immunostaining analysis showed TRP32 is cytoplasmic protein. These findings suggest that TRP32 is a novel cytoplasmic regulator of the redox state in higher eukaryotes.
Lee
K K
KK
Institute for Virus Research, Kyoto University, Kyoto 606-8507, Japan.
Murakawa
M
M
Takahashi
S
S
Tsubuki
S
S
Kawashima
S
S
Sakamaki
K
K
Yonehara
S
S
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D018922:HL-60 Cells; D006801:Humans; D019169:Jurkat Cells; D051379:Mice; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D010084:Oxidation-Reduction; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D013879:Thioredoxins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D013945:Thymoma; D014407:Tumor Cells, Cultured
D018076:DNA, Complementary; D012333:RNA, Messenger; C504766:TXNL1 protein, human; C504767:Txnl1 protein, mouse; C504765:Txnl1 protein, rat; D013879:Thioredoxins