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Caldecrin is a novel-type serine protease expressed in pancreas, but its homologue, elastase IV, is an artifact during cloning derived from caldecrin gene.
Yoshino-Yasuda, I; Kobayashi, K; Akiyama, M; Itoh, H; Tomomura, A; Saheki, T.
Afiliação
  • Yoshino-Yasuda I; Department of Biochemistry, Faculty of Medicine, Kagoshima University, Sakuragaoka 8-35-1, Kagoshima 890-8520, Japan.
J Biochem ; 123(3): 546-54, 1998 Mar.
Article em En | MEDLINE | ID: mdl-9538241
As reported previously, caldecrin, a serum calcium-decreasing factor, from pancreas was found to be a serine protease, but the proteolytic activity was not necessary for its serum calcium decreasing activity. The caldecrin cDNA encoded a protease zymogen of the chymotrypsin/elastase superfamily consisting of a signal peptide, an activation peptide and a mature enzyme. On a homology search, we found that the sequence of rat caldecrin is almost identical to that of rat elastase IV (nucleotides: 99.3%175B amino acids: 90.3%) with the exception of the central region. However, it is not known whether or not elastase IV is transcribed and translated in vivo, and has proteolytic activity. In the present study, we constructed a rat elastase IV cDNA by means of combinatorial PCR, and compared the recombinant elastase IV with the recombinant caldecrin synthesized in a baculovirus expression system. The recombinant caldecrin protein was expressed in the cells and secreted mainly into the medium. In contrast, in the case of elastase IV, no recombinant protein was immunologically or enzymatically detected in the medium, while an immunoreactive protein with much lower protease activity was found in the cells in an amount comparable to that of the caldecrin protein. Using the RT-PCR method to discriminate caldecrin mRNA from elastase IV mRNA, we detected caldecrin mRNA expression in rat pancreas, but no elastase IV mRNA expression in any tissues examined. PCR analysis of rat genomic DNA revealed the presence of caldecrin and the absence of elastase IV sequences. These results indicate that caldecrin is expressed in the pancreas, but that elastase IV is an artifact produced during cloning. Furthermore, we investigated the protein-chemical and enzymological properties of the rat and human caldecrins using their recombinant proteins. Both recombinant proteins were secreted into the medium as proforms and showed protease activity after trypsin treatment. Some differences were found in the activation process and stability between human and rat caldecrins; human caldecrin was more easily activated by trypsin, but was much more labile than rat caldecrin. Although both caldecrins were found to be chymotrypsin-type proteases, on the basis of their substrate and inhibitor specificities, they were not inhibited by TPCK, suggesting that caldecrin is a novel type of serine protease.
Assuntos
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Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Pâncreas / Serina Endopeptidases / Elastase Pancreática Limite: Animals / Humans Idioma: En Ano de publicação: 1998 Tipo de documento: Article
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Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Pâncreas / Serina Endopeptidases / Elastase Pancreática Limite: Animals / Humans Idioma: En Ano de publicação: 1998 Tipo de documento: Article