Pig leukocyte cysteine proteinase inhibitor (PLCPI), a new member of the stefin family

Lenarčič, Brigita; Ritonja, Anka; Dolenc, Iztok; Stoka, Veronika; Berbić, Selma; Pungerčar, Jože; Štrukelj, Borut; Türk, Vito

doi:10.1016/0014-5793(93)80822-c

Cited by 38 publications

(24 citation statements)

References 26 publications

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“…The porcine homologue of human stefin A, which was isolated and characterised recently [48], extends this study and provides an example of a natural mutant. It has an additional five residues at the amino terminus compared to human stefins B and A.…”

Section: Discussionmentioning

confidence: 77%

Elongation on the Amino‐terminal Part of Stefin B Decreases Inhibition of Cathepsin H

Jerala

Kroon-Zitko

Popović

et al. 1994

European Journal of Biochemistry

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Two mutants of the cysteine proteinase inhibitor, stefin B, were prepared by ligating the aminoterminal region from cystatin C and kininogen, members of two other families of cystatin superfamily. The mutant proteins were expressed in Escherichia coli and purified to homogeneity. Inhibition and kinetic constants were determined for authentic and mutated stefins against the four different cysteine proteinases, papain and human cathepsins B, L and H. Inhibition of both amino-terminal elongated stefin B mutants was decreased particularly for cathepsin H.A model of the tertiary structure of cathepsin H and its complex with stefin B was constructed. The framework for the model of cathepsin H consisted of structurally conserved regions from tertiary structures of three cysteine proteinases. Variable regions were selected from fragments of other proteins from the protein data base. We suggest that reduced binding of stefins with elongated amino termini is caused by the mini chain of cathepsin H which is probably in close proximity to the amino termini in the complexes. This mini chain is bridged to Cys214 and has already been proposed to be responsible for the aminopeptidase activity of cathepsin H. We conclude that the amino-terminal region of stefin B plays an important role in determining the strength of inhibition of cathepsin H.Cells of animals and plants contain a variety of cysteine proteinases (CP) similar to papain [I]. Mammalian cysteine proteinases include the lysosomal enzymes cathepsins B, H, L, S and C. These enzymes differ from each other in many respects, such as their pH optima or substrate specificity [2]. Some of them are true endopeptidases, while others display exclusively or additional aminopeptidase (cathepsin H) 13, 41 or carboxypeptidase activity (cathepsin B) [2]. The involvement of CPs in different physiological processes is variable as well. Some cathepsins degrade primarily proteins of the cells themselves in the process of aulophagy, while others take part in the degradation of foreign proteins in processes such as antigen processing and presentation [S -71.The inhibition of cysteine proteinases by protein inhibitors is an important mechanism for regulation of their activity [8]. The number of different cystatins (protein inhibitors which inhibit cysteine proteinases) exceeds the number of target enzymes [9]. The superfamily of cystatins of mammalian origin is usually divided into three families ; stefins, cystatins and kininogens [9, 101. In other organisms, there are more inhibitors which can be classified into those three families or form new families with intermediate properties [11]. Up to now, several cystatins have been isolated from human Knowledge on the mechanism of inhibition of cystatins is based on the crystallographic studies, where the tertiary structures of chicken cystatin [21] and human stefin B [22] were determined as representatives of two families. The mode of interaction with cysteine proteinases was first predicted from computer docking 1211 which was later confirme...

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Section: Discussionmentioning

confidence: 77%

Elongation on the Amino‐terminal Part of Stefin B Decreases Inhibition of Cathepsin H

Jerala

Kroon-Zitko

Popović

et al. 1994

European Journal of Biochemistry

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“…The cathelin protein was first isolated from porcine leucocytes and found to contain 101 amino acid residues (7). On the basis of sequence similarity, it was suggested to be an inhibitor of thiol proteases, a claim that was later retracted (8). The groups of Romeo and Zanetti and coworkers (9,10) In insects, several gene clusters for peptide antibiotics are well studied (1), but in mammals, relatively few genes for peptide antibiotics have been sequenced.…”

mentioning

confidence: 99%

Structure of the gene for porcine peptide antibiotic PR-39, a cathelin gene family member: comparative mapping of the locus for the human peptide antibiotic FALL-39.

Guðmundsson

Magnússon

Chowdhary

et al. 1995

Proc. Natl. Acad. Sci. U.S.A.

116

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PR-39 is a porcine 39-aa peptide antibiotic composed of 49% proline and 24% arginine, with an activity against Gram-negative bacteria comparable to that of tetracycline. In Escherichia coli, it inhibits DNA and protein synthesis. PR-39 was originally isolated from pig small intestine, but subsequent cDNA cloning showed that the gene is expressed in the bone marrow. The open reading frame of the clone showed that PR-39 is made as 173-aa precursor whose proregion belongs to the cathelin family. The PR39 gene, which is rather compact and spans only 1784 bp has now been sequenced. The coding information is split into four exons. The first exon contains the signal sequence of 29 residues and the first 37 residues of the cathelin propart. Exons 2 and 3 contain only cathelin information, while exon 4 codes for the four C-terminal cathelin residues and the mature PR-39 peptide extended by three residues. The sequenced upstream region (1183 bp) contains four potential recognition sites for NF-IL6 and three for APRF, transcription factors known to regulate genes for both cytokines and acute phase response factors. Genomic hybridizations revealed a fairly high level of restriction fragment length polymorphism and indicated that there are at least two copies of the PR39 gene in the pig genome. PR39 was mapped to pig chromosome 13 by linkage and in situ hybridization mapping. The gene for the human peptide antibiotic FALL-39 (also a member of the cathelin family) was mapped to human chromosome 3, which is homologous to pig chromosome 13.

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“…The C-terminal peptide is released in mature form after proteolytic processing and is highly variable between members of this same family (41). Initially, cathelin was identified as a cysteine protease inhibitor but this activity was later attributed to a protein which was inadvertently copurified with cathelin (33). The function of the proregion within the cathelicidins has been theorized to neutralize the charged C-terminal portion to prevent antibiotic effects of this region before processing, target the protein to the appropriate organelle, assist in folding of the mature protein, or serve a novel function yet to be identified (42).…”

Section: Discussionmentioning

confidence: 99%

“…These unidentified proteins do not represent modified forms of the ␥ or subunits but each is labeled with cell surface iodination. We have now identified one of these unique Fc␥R-associated proteins and have also demonstrated an association between Fc␥RIIIa␣ and the 15-kDa protein that contains homology to cathelin, a protein of undefined function identified initially in porcine leukocytes (33). A cathelin domain is contained within the family of antimicrobial proteins known as cathelicidins.…”

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Identification of a Novel FcγRIIIaα-Associated Molecule That Contains Significant Homology to Porcine Cathelin

Sweeney

Kim

2004

The Journal of Immunology

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The following studies are the first to demonstrate the association of porcine FcγRIIIaα with a molecule that contains significant homology to the cathelin family of antimicrobial proteins. We performed immunoprecipitation of the porcine FcγRIIIaα multisubunit complex from Brij 96 lysates of polymorphonuclear leukocytes using the G7 mAb, which binds to FcγRIIIaα on the surface of porcine NK cells and phagocytes. Previous results indicate that the transmembrane α subunit of the FcγRIIIa complex is associated with the γ subunit on the surface of porcine polymorphonuclear leukocytes and with several other unique proteins that surface iodinate and migrate at ∼15, 20, and 25 kDa when analyzed by reducing SDS-PAGE. Through characterization of the porcine FcγRIIIa complex, we identified the 15-kDa molecule as a unique FcγR-associated protein that has not been described in other systems. We now report an association between FcγRIIIaα and a 15-kDa molecule that shares homology to cathelin, a protein of undetermined function initially identified in porcine leukocytes. A domain with a high degree of homology to cathelin is found in the proregions of a family of antibiotic proteins referred to as cathelicidins. The results of our studies indicate the presence of a novel FcγRIIIa complex in the porcine system, and may provide new insights into the function of this antimicrobial protein homologue in relation to the variety of responses mediated through FcγRs.

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Pig leukocyte cysteine proteinase inhibitor (PLCPI), a new member of the stefin family

Cited by 38 publications

References 26 publications

Elongation on the Amino‐terminal Part of Stefin B Decreases Inhibition of Cathepsin H

Elongation on the Amino‐terminal Part of Stefin B Decreases Inhibition of Cathepsin H

Structure of the gene for porcine peptide antibiotic PR-39, a cathelin gene family member: comparative mapping of the locus for the human peptide antibiotic FALL-39.

Identification of a Novel FcγRIIIaα-Associated Molecule That Contains Significant Homology to Porcine Cathelin

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