Molecular cloning and sequence analysis of human preprocathepsin C

Pariš, Alenka; Štrukelj, Borut; Pungerčar, Jože; Renko, M.; Dolenc, Iztok; Türk, Vito

doi:10.1016/0014-5793(95)00777-7

Cited by 79 publications

(51 citation statements)

References 27 publications

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“…Nevertheless, the value is comparable to those of other chicken cystatin/cathepsin interactions (B. Turk, unpublished results). This difference could be explained by Trp26Tyr and Trp69Tyr substitutions (papain numbering) in cathepsin C [29].…”

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confidence: 98%

Interaction of human cathepsin C with chicken cystatin

et al. 1996

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Cathepsin C was purified from human spleen by a rapid procedure, which included homogenization, ammonium sulfate precipitation, gel filtration on Sephacryl S-200 and fmally affinity chromatography on chicken cystatin-Sepharose. The interaction between cathepsin C and chicken cystatin was further characterized. It was found to be accompanied by a maximum decrease in fluorescence emission intensity at 330 nm. Fluorescence titration showed that human catbepsin C can bind four chicken cystatin molecules. The 4:1 binding stoichiometry was confirmed by titration monitored by the loss of enzyme activity. A non-competitive-competitive type of inhibition was determined from a double-reciprocal Lineweaver-Burk plot with a Kj value of 0.22 nM for the non-competitive inhibition.

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confidence: 98%

Interaction of human cathepsin C with chicken cystatin

et al. 1996

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“…A human and a rat DPP-I cDNA have recently been cloned (9,10), but the reported sequences contained only a portion of the 5Ј-untranslated regions (UTRs). Moreover, information on gene expression and regulation has not been reported.…”

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Human Dipeptidyl-peptidase I

Rao

Hoidal

1997

Journal of Biological Chemistry

146

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Dipeptidyl-peptidase I, a lysosomal cysteine proteinase, is important in intracellular degradation of proteins and appears to be a central coordinator for activation of many serine proteinases in immune/inflammatory cells. Little is known about the molecular genetics of the enzyme. In the present investigation the gene for dipeptidyl-peptidase I was cloned and characterized. The gene spans approximately 3.5 kilobases and consists of two exons and one intron. The genomic organization is distinct from the complex structures of the other members of the papain-type cysteine proteinase family. By fluorescence in situ hybridization, the gene was mapped to chromosomal region 11q14.1-q14.3. Analysis of the sequenced 5 -flanking region revealed no classical TATA or CCAAT box in the GC-rich region upstream of cap site. A number of possible regulatory elements that could account for tissue-specific expression were identified. Northern analyses demonstrated that the dipeptidyl-peptidase I message is expressed at high levels in lung, kidney, and placenta, at moderate to low levels in many organs, and at barely detectable levels in the brain, suggesting tissue-specific regulation. Among immune/inflammatory cells, the message is expressed at high levels in polymorphonuclear leukocytes and alveolar macrophages and their precursor cells. Treatment of lymphocytes with interleukin-2 resulted in a significant increase in dipeptidyl-peptidase I mRNA levels, suggesting that this gene is subjected to transcriptional regulation. The results provide initial insights into the molecular basis for the regulation of human dipeptidylpeptidase I.

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“…Antibodies directed against bovine cathepsin S were purified from a rabbit antiserum, obtained by immunization with cathepsin S from bovine spleen . A human ileum &I1 cDNA library was constructed as described (Paris et al, 1995).…”

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Folding and Activation of Human Procathepsin S from Inclusion Bodies Produced in Escherichia coli

Kopitar

Dolinar

Štrukelj

et al. 1996

European Journal of Biochemistry

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Human procathepsin S was produced in the form of insoluble inclusion bodies in Escherichia coli using an inducible T7-based expression system. After cell disruption, the dissolved inclusion body proteins were S-sulphonated with 2-nitro-5-thiosulphobenzoate and purified by gel filtration. Recombinant procathepsin S was renatured at pH 7.6 by a two-step dilution which significantly increased the yield of production compared to single-step dilution. The proenzyme was autocatalytically processed to active cathepsin S at pH 4.5 in the presence of an excess of cysteine and catalytic amounts of dextran sulphate. Most of the loss of procathepsin S occurred during folding, probably because of aggregation. Concentrations lower than 20 pg/ml of procathepsin S were necessary to minimise such aggregation. The recombinant cathepsin S was catalytically active on fluorogenic substrates and had kinetic properties similar to those of recombinant enzyme produced in yeast. The expression, renaturation, and activation procedures used enable the production of up to 2 mg of catalytically active recombinant human cathepsin S/1 fermentation broth.Keywords: cathepsin S ; Escherichia coli ; inclusion bodies ; proenzyme ; renaturation ; processing.Cysteine proteinases, including cathepsin S, are capable of degrading extracellular matrix in various diseases (Sloane et al., 1990;Lah et al., 1992; GabrijeltiE et al., 1992). The degradation is thought to occur in the extracellular environment where the pH is neutral; among cysteine proteinases, only cathepsin S is active at this pH. The highest expression of human cathepsin S mRNA was found in lungs, heart muscle, and spleen, with a significantly lower expression in other tissues including brain (Shi et al., 1994), where the expression of rat cathepsin S is high (Petanceska and Devi, 1992). Very recently, it has been reported that cathepsin S may play a role in pathogenesis of the cerebral lesions of Alzheimer's disease or represent a response to their formation. It was suggested that cathepsin S could be partly involved in processing p-amyloid precursor protein to amyloid-p in the amyloid plaques (Lemere et al., 1995).Cathepsin S is a lysosomal cysteine proteinase (Turk and Bode, 1993). In contrast to other cathepsins, cathepsin S is present exclusively as a single-chain proteinase (Ritonja et al., 1991 ;Wiederanders et al., 1991) and is not glycosylated, although it contains a potential glycosylation site on its propeptide (Shi et al., 1992;Wiederanders et al., 1992). In contrast to cathepsins B and L, cathepsin S is both stable and active at neutral pH. It has a high endopeptidase activity against different proteins including elastin and collagen. Its substrate specificity shows some similarities with cathepsin L, but it differs distinctly in its S, subsite specificity (Xin et al., 1992).Human cathepsin S cDNA encodes a preproprotein of 331 amino acid residues, whereas the mature enzyme contains 217 residues (Shi et al., 1992;Wiederanders et al., 1992).The cDNA for human preprocathepsin S h...

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Molecular cloning and sequence analysis of human preprocathepsin C

Cited by 79 publications

References 27 publications

Interaction of human cathepsin C with chicken cystatin

Interaction of human cathepsin C with chicken cystatin

Human Dipeptidyl-peptidase I

Folding and Activation of Human Procathepsin S from Inclusion Bodies Produced in Escherichia coli

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