Isolation of a human cDNA for .alpha.2-thiol proteinase inhibitor and its identity with low molecular weight kininogen

Abstract

A lambda gt11 cDNA library containing DNA inserts prepared from human liver mRNA has been screened with an antibody to human alpha 2-thiol proteinase inhibitor that was isolated from fresh plasma. Eighteen positive clones were isolated from one million phage, and each was plaque purified. The cDNA insert of one of these phage was sequenced and shown to code for alpha 2-thiol proteinase inhibitor as identified by a partial amino acid sequence of the light chain of alpha 2-thiol proteinase inhibitor. This cDNA insert contained 1529 base pairs coding for the complete alpha 2-thiol proteinase inhibitor. It included 45 base pairs of 5' noncoding sequence, 1281 base pairs that code for pre alpha 2-thiol proteinase inhibitor, a stop codon, 160 base pairs of 3' noncoding sequence, and 40 base pairs of poly(A) tail. The noncoding sequence on the 3' end contained a potential recognition site (AATAAA) for processing and polyadenylation of precursor messenger RNA. The amino acid sequence of alpha 2-thiol proteinase inhibitor deduced from the cDNA showed a striking similarity (overall homology at 74%) to that of bovine low molecular weight (LMW) kininogen, including two internally repeated sequences and a nonapeptide sequence of bradykinin. These data clearly indicated that alpha 2-thiol proteinase inhibitor and LMW kininogen are identical. This was further supported by immunological cross-reactivity between alpha 2-thiol proteinase inhibitor and LMW kininogen.(ABSTRACT TRUNCATED AT 250 WORDS)

Friends and relations of the cystatin superfamily—new members and their evolution

“…Cystatin-like sequences identified in the kininogens (Ohkubo et al, 1984) were found to be present as sequence duplications. Subsequently, it was proposed that there were three cystatin-like repeats in the kininogen sequence, the amino terminal repeat being Reprint requests to: K.M Dziegielewska, Department of Anatomy and Physiology, University of Tasmania, GPO Box 252-24, Hobart, Tasmania 7001, Australia.…”

“…The concept of a cystatin "superfamily" emerged in the early 1980s, precipitated by an observation that multiple cystatin-like sequences were present in the kininogens and that the stefins were related to both the cystatins and the repeats in the kininogens (Ohkubo et al, 1984). The cystatins are a family of cysteine protease inhibitors with homology to chicken cystatin (Barrett, 1981).…”

“…Cystatin superfamily members have been found throughout evolution and members of each family of the superfamily are present in mammals today. In this review, the new and older, established members of the family are arranged into a possible evolutionary order, based on sequence homology and functional similarities.Keywords: cystatins; cysteine protease inhibitors; evolution; fetuins; kininogens; TGF-/3 receptor I1The concept of a cystatin "superfamily" emerged in the early 1980s, precipitated by an observation that multiple cystatin-like sequences were present in the kininogens and that the stefins were related to both the cystatins and the repeats in the kininogens (Ohkubo et al, 1984). The cystatins are a family of cysteine protease inhibitors with homology to chicken cystatin (Barrett, 1981).…”

“…Their structural genes comprise three exons of characteristic size (e.g., Huh et al, 1995) (Fig. I), and a cluster of these genes is located on human chromosome 20 (e.g., Thiesse et al, 1994) although cystatin genes are also found on other chromosomes (e.g., Pennachio et al, 1996).Cystatin-like sequences identified in the kininogens (Ohkubo et al, 1984) were found to be present as sequence duplications. Subsequently, it was proposed that there were three cystatin-like repeats in the kininogen sequence, the amino terminal repeat being Reprint requests to: K.M Dziegielewska, Department of Anatomy and Physiology, University of Tasmania, GPO Box 252-24, Hobart, Tasmania 7001, Australia.…”

“…These two hairpin loops and the largely unstructured amino terminus of the protein form a wedge that fits into the protease (Bode et al, 1988;Stubbs et al, 1990;Machleidt et al, 1995; Auerswald et al, 1996). In the fetuins and HRGs, these sequences have been extensively altered, making it unlikely that they are viable cysteine protease inhibitors.Conserved sequences around the cysteine at the carboxylterminus of the fetuins, HMW kininogen, and HRG As Elzanowski et al (Elzanowski et al, 1988) noted, the sequence immediately surrounding the last cysteine residue, pro-pro-_cys-pro-gly-arg in human fetuin (Gejyo et al, 1983;Lee et al, 1987) shares significant homology with the same region of human prolys-*-pro-gly-arg (Ohkubo et al, 1984;Kitamura et al, 1985;Salvesen et al, 1986;Kellermann et al, 1986Kellermann et al, , 1987, and bovine pro-lys-%-pro-ser-arg Kitamura et al, 1983;Sueyoshi et al, 1984Sueyoshi et al, , 1985 high-molecular weight kininogen, suggesting a possible common origin of the carboxyl-terminal regions of the two proteins. Furthermore, the sequence around the last cysteine in bovine glu-ser-cys-pro-gly-thr (Sorensen et al, 1993) and human glu-ser-cys-pro-gly-lys (Koide et al, 1986) histidine-rich glycoprotein is also similar.…”