Kx. Chai et al., KALLISTATIN - A NOVEL HUMAN SERINE PROTEINASE-INHIBITOR - MOLECULAR-CLONING, TISSUE DISTRIBUTION, AND EXPRESSION IN ESCHERICHIA-COLI, The Journal of biological chemistry, 268(32), 1993, pp. 24498-24505
We have recently purified a novel human serine proteinase inhibitor (s
erpin), designated as kallistatin, which binds to tissue kallikrein an
d inhibits kallikrein's kininogenase and amidolytic activities. In the
present studies, we have cloned a full-length cDNA encoding kallistat
in from human liver RNA by the polymerase chain reaction. The cDNA is
1284 base pairs in length and encodes 427 amino acid residues, includi
ng a 26-residue signal peptide and a 401-residue mature peptide. The t
ranslated amino acid sequence of kallistatin matches with the protein
sequence and shares 44-46% sequence identity with human alpha1-antichy
motrypsin, protein C inhibitor, corticosteroid-binding globulin, alpha
1-antitrypsin, thyroxin-binding globulin, and rat kallikrein-binding p
rotein. Kallistatin is a new member of the serpin superfamily with a u
nique reactive site P1-P1' of Phe-Ser. Four potential glycosylation si
tes are found in the translated amino acid sequence of kallistatin. In
a Southern blot analysis following reverse transcription and polymera
se chain reaction, kallistatin was found to be expressed in human live
r, stomach, pancreas, kidney, aorta, testes, prostate, artery, atrium,
ventricle, lung, renal proximal tubular cell, and a colonic carcinoma
cell line T84. A genomic Southern blot using the full-length kallista
tin cDNA probe revealed simple banding patterns suggesting the gene en
coding kallistatin is single-copied. The kallistatin cDNA encoding the
mature peptide was expressed in Escherichia coli. The recombinant kal
listatin forms an SDS-stable complex with I-125-human tissue kallikrei
n and has a molecular mass of 40 kDa. The cloning of human kallistatin
cDNA established the identity of the novel kallikrein inhibitor and i
ts expression in a functional form in E. coli provides means for study
ing its structure-function relationship through protein engineering.