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Human cystatin D: cDNA cloning, characterization of the E. coli expressed inhibitor, and identification of the native protein in saliva

Freije, José P; Balbin, Milagros; Abrahamson, Magnus LU ; Velasco, Gloria; Dalboge, Henrik; Grubb, Anders LU and Lopez-Otin, Carlos (1993) In Journal of Biological Chemistry 268(21). p.15737-15744
Abstract
A cDNA coding for cystatin D, a human member of the cystatin protein family, has been cloned after specific amplification of reverse- transcribed parotid gland RNA. After replacing the segment encoding the putative 20-residue signal peptide with one encoding the Escherichia coli OmpA leader sequence, the cDNA was expressed in E. coli. The isolated recombinant protein exhibited Ki values of 1.2 nM and > 1 microM for papain and cathepsin B, respectively. An antiserum raised against recombinant cystatin D recognized a protein in human saliva with electrophoretical mobility identical to that of the recombinant protein. Immunoenzymatic analysis revealed that this cysteine proteinase inhibitor is present in human saliva and tears at... (More)
A cDNA coding for cystatin D, a human member of the cystatin protein family, has been cloned after specific amplification of reverse- transcribed parotid gland RNA. After replacing the segment encoding the putative 20-residue signal peptide with one encoding the Escherichia coli OmpA leader sequence, the cDNA was expressed in E. coli. The isolated recombinant protein exhibited Ki values of 1.2 nM and > 1 microM for papain and cathepsin B, respectively. An antiserum raised against recombinant cystatin D recognized a protein in human saliva with electrophoretical mobility identical to that of the recombinant protein. Immunoenzymatic analysis revealed that this cysteine proteinase inhibitor is present in human saliva and tears at concentrations of 3.8 and 0.5 mg/liter, respectively, while it was not detected in seminal plasma, blood plasma, milk, or cerebrospinal fluid. Cystatin D purified from human saliva by immunosorption displayed a heterogeneous N-terminal end, with sequences starting at residues 5, 7, 9, and 11 of the predicted N-terminal portion of the mature protein. On the basis of structural and functional properties, cystatin D represents a novel cysteine proteinase inhibitor possibly playing a protective role against proteinases present in the oral cavity. (Less)
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author
organization
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type
Contribution to journal
publication status
published
subject
in
Journal of Biological Chemistry
volume
268
issue
21
pages
15737 - 15744
publisher
ASBMB
ISSN
1083-351X
language
English
LU publication?
yes
id
23272562-6fe1-4bf2-9eeb-f97b7213ac33 (old id 1107645)
alternative location
http://www.jbc.org/cgi/reprint/268/21/15737
date added to LUP
2008-07-31 14:06:59
date last changed
2016-04-15 18:55:02
@article{23272562-6fe1-4bf2-9eeb-f97b7213ac33,
  abstract     = {A cDNA coding for cystatin D, a human member of the cystatin protein family, has been cloned after specific amplification of reverse- transcribed parotid gland RNA. After replacing the segment encoding the putative 20-residue signal peptide with one encoding the Escherichia coli OmpA leader sequence, the cDNA was expressed in E. coli. The isolated recombinant protein exhibited Ki values of 1.2 nM and > 1 microM for papain and cathepsin B, respectively. An antiserum raised against recombinant cystatin D recognized a protein in human saliva with electrophoretical mobility identical to that of the recombinant protein. Immunoenzymatic analysis revealed that this cysteine proteinase inhibitor is present in human saliva and tears at concentrations of 3.8 and 0.5 mg/liter, respectively, while it was not detected in seminal plasma, blood plasma, milk, or cerebrospinal fluid. Cystatin D purified from human saliva by immunosorption displayed a heterogeneous N-terminal end, with sequences starting at residues 5, 7, 9, and 11 of the predicted N-terminal portion of the mature protein. On the basis of structural and functional properties, cystatin D represents a novel cysteine proteinase inhibitor possibly playing a protective role against proteinases present in the oral cavity.},
  author       = {Freije, José P and Balbin, Milagros and Abrahamson, Magnus and Velasco, Gloria and Dalboge, Henrik and Grubb, Anders and Lopez-Otin, Carlos},
  issn         = {1083-351X},
  language     = {eng},
  number       = {21},
  pages        = {15737--15744},
  publisher    = {ASBMB},
  series       = {Journal of Biological Chemistry},
  title        = {Human cystatin D: cDNA cloning, characterization of the E. coli expressed inhibitor, and identification of the native protein in saliva},
  volume       = {268},
  year         = {1993},
}