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ABSTRACT: Senescence marker protein-30 (SMP30), composed of 299 amino acids, has an approximate molecular mass of 32-34 kDa and has a pI 4.9 in charge. The amino acid alignment from various animal species revealed a highly conserved structure. SMP30 has an enzyme activity hydrolyzing sarin, soman, and tabun, known as lethal toxic nerve chemicals. We analyzed the organophosphatase activity of SMP30 using DFP as a substrate. This DFPase activity is revealed in a dose-dependent manner in the presence of magnesium ions. We investigated the intracellular localization of SMP30. It is localized in both the cytoplasm and nucleus. To confirm the presence of SMP30 in the nucleus, we prepared nuclear and cytoplasmic extracts from isolated cultured hepatocytes. Western blotting showed that SMP30 was detected in both extracts. Because the expression is reduced by carbon tetrachloride, one can speculate that the expression is modulated by oxidative stress increased with aging.
Annals of the New York Academy of Sciences 07/2004; 1019:360-4. · 3.15 Impact Factor
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ABSTRACT: A mouse strain lacking SMP30 can be regarded as a strain showing ultimate decrease of the SMP30 molecule. After three months of age, SMP30-KO mice had an increased mortality rate, compared with the SMP30-WT mice, all of which remained alive. Electron microscopic observation of the hepatocytes from 12-month-old SMP30-KO mice revealed many empty vacuoles, presumably lipid droplets, abnormally enlarged mitochondria with indistinct cristae, and exceptionally large lysosomes filled with electron-dense bodies. The total hepatic triglyceride concentration of SMP30-KO mice was approximately 3.6-fold higher than that of the age-matched wild type. Similarly, the total hepatic cholesterol of SMP30-KO mice reached an approximate 3.3-fold greater value than that of the comparative group. Total hepatic phospholipids of SMP30-KO mice achieved an approximately 3.7-fold higher level compared with that of the wild-type mice. The cells from SMP30-KO mice were sensitive to apoptotic reagents. Those results supported the idea that SMP30 has an antiapoptotic function with wide spectrum. These findings indicate that SMP30-KO mice are highly susceptible to various harmful reagents. This strain might be a useful tool for aging and biological monitoring.
Annals of the New York Academy of Sciences 07/2004; 1019:383-7. · 3.15 Impact Factor
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ABSTRACT: Peptidylarginine deiminases (PADs) are posttranslational modification enzymes that convert protein arginine to citrulline residues in a calcium ion-dependent manner. Rodents have four isoforms of PAD (types I, II, III, and IV), each of which is distinct in substrate and tissue specificity. In fact, the only tissue in which all four PAD mRNAs have been detected is the epidermis. In this study, we found PAD activity in HSC-1 human cutaneous squamous carcinoma cells in vitro, and this activity increased during cultivation. Using a homology-based strategy, we cloned a full-length cDNA encoding human PAD type II. The cDNA was 2348 bp long and encoded a 665-amino-acid sequence with a predicted molecular mass of 75 kDa. The predicted protein shared 93% identity with the rat and mouse PAD type II sequence. Alignment of the amino acid sequences from both species revealed notable conservation in the C-terminal region, suggesting the presence of a functional region such as an enzyme catalytic site and/or a calcium-binding domain. Gene organization analysis established that human PAD type II on chromosome 1p35.2-p35.21 spanned more than 50 kb and contained 16 exons and 15 introns. A recombinant PAD protein subsequently produced in Escherichia coli proved to be enzymatically active, with substrate specificities similar to those of the rat PAD type II. In an immunohistochemical study of human skin, the type II enzyme was expressed by all the living epidermal layers, suggesting that PAD type II is functionally important during terminal differentiation of epidermal keratinocytes.
Archives of Biochemistry and Biophysics 12/2002; 407(1):25-31. · 2.93 Impact Factor
