Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Cystatins”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 865 records · Page 48Linked to original sources
PubMed1996

Cystatin-like domain of mouse countertrypin, a member of mammalian fetuin family, is responsible for the inhibition of trypsin. Evidence from site-directed mutagenesis.

Members of mammalian fetuin family, such as human alpha 2 HS glycoprotein and bovine fetuin, consist of three domains, two tandemly arranged cystatin-like domains and an unrelated domain, but they have no inhibitory activity against cysteine proteinases. We found that countertrypin, a novel trypsin inhibitor, is mouse counterpart of human alpha 2 HS glycoprotein, and that human alpha 2 HS glycoprotein and bovine fetuin which were prepared without use of ethanol are capable of inhibiting trypsin (Yamamoto, K. and Sinohara, H. (1993) J. Biol. Chem, 268, 17750-17753). In the present study, cDNA encoding countertrypin was isolated and sequenced, and evidence is presented, based on the site-directed mutagenesis, that lysine-231 in the second cystatin domain is the P1 site for trypsin inhibition.

Amino Acid Sequence↗
PubMed2004

The barley cystatin gene (Icy) is regulated by DOF transcription factors in aleurone cells upon germination.

A genomic clone (Icy gene) encoding a barley cystatin has been characterized. The gene contains one intron interrupting its ORF that encodes a protein of 107 amino acid residues. A DNA fragment of -1058 bp upstream of the ATG translation initiation codon has been sequenced and several promoter deletions fused to the beta-glucuronidase (GUS; uidA gene) reporter gene obtained. Transient expression assays in different barley tissues have indicated that a -631 bp promoter fragment was sufficient for full activity. In bombarded barley aleurone layers the GUS-driven expression by this promoter is repressed by GA(3) incubation, as is the accumulation of the Icy transcripts detected. A spatial and temporal location of cystatin transcripts by in situ hybridization techniques indicates that this gene is ubiquitously expressed and its transcripts are particularly abundant in leaves and roots, and in seeds, both during development and upon germination. Two DOF transcription factors, SAD and BPBF, previously described as involved in gene expression regulation in seeds, interact specifically in vitro with oligonucleotides containing DOF binding-sites derived from the Icy gene promoter. Transient expression experiments in co-bombarded aleurone layers demonstrate that BPBF strongly represses transcription of the native Icy promoter, even when co-transfected with SAD that behaves as an activator in this in vivo system.

Amino Acid Sequence↗
PubMed1994

Corn cystatin I expressed in Escherichia coli: investigation of its inhibitory profile and occurrence in corn kernels.

Corn cystatin I was expressed in Escherichia coli as a mature protein. It was purified by gel filtration on Sephadex G-50, ion-exchange HPLC, and reversed-phase HPLC. The purified protein showed strong inhibitory activities against papain (Ki: 3.7 x 10(-8) M), and cathepsins H (Ki: 5.7 x 10(-9) M) and L (Ki: 1.7 x 10(-8) M), whereas it inhibited cathepsin B to a lesser extent (Ki: 2.9 x 10(-7) M). Western blot analysis using antibody raised against corn cystatin I revealed that in the corn kernel, the protein occurs with a molecular mass of approximately 13 kDa. Localization in the aleurone layer and embryo of the corn kernel was shown by immunostaining microscopy.

Cloning, Molecular↗
PubMed1999

Immunolocalization of CRES (Cystatin-related epididymal spermatogenic) protein in the acrosomes of mouse spermatozoa.

The CRES (cystatin-related epididymal spermatogenic) protein is a member of the cystatin superfamily of cysteine protease inhibitors and exhibits highly restricted expression in the reproductive tract. We have previously shown that CRES protein is present in elongating spermatids in the testis and is synthesized and secreted by the proximal caput epididymal epithelium. The presence of CRES protein in developing germ cells and in the luminal fluid surrounding maturing spermatozoa prompted us to examine whether CRES protein is associated with spermatozoa. In the studies presented, indirect immunofluorescence, immunogold electron microscopy, and Western blot analysis demonstrated that CRES protein is localized in sperm acrosomes and is released during the acrosome reaction. Interestingly, while the 19- and 14-kDa CRES proteins were present in testicular and proximal caput epididymal spermatozoa, the 14-kDa CRES protein was the predominant form present in mid-caput to cauda epididymal spermatozoa. Furthermore, following the ionophore-induced acrosome reaction, CRES protein localization was similar to that of proacrosin/acrosin in that it was detected in the soluble fraction as well as associated with the acrosome-reacted spermatozoa. The presence of CRES protein in the sperm acrosome, a site of high hydrolytic and proteolytic activity, suggests that CRES may play a role in the regulation of intraacrosomal protein processing or may be involved in fertilization.

Acrosin↗
PubMed1993

Proteolysis of the 85-kilodalton crystalline cysteine proteinase inhibitor from potato releases functional cystatin domains.

The protein crystals found in potato (Solanum tuberosum L.) tuber cells consist of a single 85-kD polypeptide. This polypeptide is an inhibitor of papain and other cysteine proteinases and is capable of binding several proteinase molecules simultaneously (P. Rodis, J.E. Hoff [1984] Plant Physiol 74: 907-911). We have characterized this unusual inhibitor in more detail. Titrations of papain activity with the potato papain inhibitor showed that there are eight papain binding sites per inhibitor molecule. The inhibition constant (Ki) value for papain inhibition was 0.1 nM. Treatment of the inhibitor with trypsin resulted in fragmentation of the 85-kD polypeptide into a 32-kD polypeptide and five 10-kD polypeptides. The 32-kD and 10-kD fragments all retained the ability to potently inhibit papain (Ki values against papain were 0.5 and 0.7 nM, respectively) and the molar stoichiometries of papain binding were 2 to 3:1 and 1:1, respectively. Other nonspecific proteinases such as chymotrypsin, subtilisin Carlsberg, thermolysin, and proteinase K also cleaved the 85-kD inhibitor polypeptide into functional 22-kD and several 10-kD fragments. The fragments obtained by digestion of the potato papain inhibitor with trypsin were purified by reverse-phase high-performance liquid chromatography, and the N-terminal amino acid sequence was obtained for each fragment. Comparison of these sequences showed that the fragments shared a high degree of homology but were not identical. The sequences were homologous to the N termini of members of the cystatin superfamily of cysteine proteinase inhibitors. Therefore, the inhibitor appears to comprise eight tandem cystatin domains linked by preteolytically sensitive junctions. We have called the inhibitor potato multicystatin (PMC). By immunoblot analysis and measurement of papain inhibitory activity, PMC was found at high levels in potato leaves (up to 0.6 microgram/g fresh weight tissue), where it accumulated under conditions that induce the accumulation of other proteinase inhibitors linked to plant defense. PMC may have a similar defensive role, for example in protecting the plant from phytophagous insects that utilize cysteine proteinases for dietary protein digestion.

Amino Acid Sequence↗
PubMed1992

Candida albicans produces a cystatin-type cysteine proteinase inhibitor.

A cysteine proteinase inhibitor was found in culture media of Candida albicans. Purification to homogeneity of the inhibitor was performed by carboxymethyl-papain-Sepharose affinity, DE-52 ion-exchange, and reverse-phase high performance liquid chromatographies. The purified inhibitor had an M(r) of 15 kDa and a pI of 4.9. It was more stable to heat and pH than most proteins. The N-terminal sequence of the first 30 residues demonstrated high similarity with that of human cystatin A. Thus, C. albicans cysteine proteinase inhibitor seems to belong to the cystatin superfamily. The inhibitor activity of the yeast cellular form was 4.0 times higher than that of the hyphal cellular form in 7-day culture media. It is suggested that the inhibitor has regulatory functions similar to those of its counterpart proteinases in the invasion of host cells.

Candida albicans↗
PubMed1999

Cystatin-related epididymal spermatogenic protein colocalizes with luteinizing hormone-beta protein in mouse anterior pituitary gonadotropes.

The CRES (cystatin-related epididymal spermatogenic) protein, a member of the cystatin superfamily of cysteine protease inhibitors, exhibits highly restricted expression in the mouse testis and epididymis, suggesting roles in reproduction. Considering the well-established relationship that exists between the gonads and the neuroendocrine system, the present studies were undertaken to determine whether the CRES messenger RNA and protein are expressed in the anterior pituitary gland and, if so, whether the expression is regulated by hormones. RT-PCR analysis of whole pituitary gland RNA preparations, and Northern blot analyses of pituitary gland cell lines, demonstrated that the CRES gene is expressed in the male and female anterior pituitary gland gonadotropes. Furthermore, Western blot analysis demonstrated that CRES protein was present in whole mouse pituitary glands and was synthesized and secreted by the LbetaT2 gonadotrope cell line. Interestingly, whereas the predominant CRES proteins present in epididymal lysates, LbetaT2 secretory granules, and whole pituitary gland lysates were 19 and 14 kDa, the predominant CRES proteins present in the cell culture conditioned media were 17 and 12 kDa. Deglycosylation studies revealed that the higher-molecular-mass CRES proteins (19 and 17 kDa) were the result of N-linked glycosylation, caused by the presence of high mannose residues. Double-label immunofluorescence and confocal microscopic analysis of male and female mouse pituitary gland tissue confirmed the RNA studies and showed that CRES protein colocalized with LHbeta protein in the gonadotropes. Finally, gonadectomy and hormone replacement studies suggest that CRES protein in the gonadotropes is hormonally regulated. These studies suggest that CRES protein may perform a role in the gonadotrope-mediated control of reproduction.

Animals↗
PubMed1992

Cystatin mimicry by synthetic peptides.

Synthetic peptides which tentatively mimic the cystatin inhibitory surface were used to study the mechanism of inhibition of cysteine proteinases by their natural inhibitors. The inhibitory properties of these peptides depend mainly on the presence of the QxVxG consensus sequence. N and C-terminal peptide derivatives bearing large hydrophobic groups showed dramatically improved inhibition. Molecular dynamic studies after energy minimization showed that the non covalent interaction between these hydrophobic groups induced the formation of a loop structure which probably favours inhibition. Antibodies were raised against one of these peptides, which recognized kininogens in the serum of all mammal species tested, but not cystatins from family two.

Animals↗
PubMed1995

Regulation of the activity of lysosomal cysteine proteinases by pH-induced inactivation and/or endogenous protein inhibitors, cystatins.

The kinetics of pH-induced inactivation of human cathepsins B and L was studied by conventional and stopped-flow methods. The inactivation of both enzymes was found to be an irreversible, first-order process. The inactivation rate constants increased exponentially with pH for both enzymes. From log kinac vs pH plots, 3.0 and 1.7 protons were calculated to be desorbed for pH-induced inactivation of cathepsins L and B. Cathepsin B was thus substantially more stable than cathepsin L (approximately 15-fold at pH 7.0 and 37 degrees C). Cathepsin B was efficiently inhibited by cystatin C at pH 7.4, whereas the inhibition by stefin B and high molecular weight kininogen was only moderate. In contrast, cathepsin L was efficiently inhibited by both chicken cystatin and stefin B at this pH kass approximately 3.3 x 10(7) m-1 s-1).

Animals↗
PubMed2002

Affinity of chicken cystatin for S-(carbamoylmethyl)-papain, measured by fluorescence at acidic pH.

We have characterized the interaction between S-(carbamoylmethyl)-papain and chicken cystatin by means of fluorescence titration at acidic pH. The calculated binding constant was 32 x 10(6), which is less than the tight affinity at pH 7.4 (approximately 7 x 10(10)). Results have been discussed in terms of the burial of Trp-104 of cystatin and the possible role of the charge of the carbamoylmethyl moiety.

Animals↗
PubMed1997

Genomic structure of human cystatin A.

Cystatin A is a cysteine proteinase inhibitor with a molecular mass of 11 kDa, and is located mainly in the keratohyaline granules of the stratum granulosum and the cornified envelope of the stratum corneum in the epidermis. In this study, we demonstrated the genomic structure of this proteinase inhibitor in which there were three exons of 111 bp, 102 bp and 226 bp in length, while the lengths of the 1st and 2nd intron were approximately 14 Kbp and 4 Kbp, respectively. The conserved sequence of QVVAG was encoded in the 2nd exon and was not inserted by any introns. There were binding sites for SP-1 and AP-2 in the promoter region and an AP-1 binding site in the 1st intron. The successful amplification of each exon of cystatin A may possibly contribute to the detection of the genomic abnormality of some skin disorders e.g. keratinization disorder, chronic bacterial infection or photophobia.

Amino Acid Sequence↗
PubMed2000

IgM and IgG cystatin capture enzyme linked immunosorbent assays: a tool for serodiagnosis and assessment of cure of acute fascioliasis after triclabendazole (TCZ; Fasinex) therapy.

This study included 71 individuals: 25 patients with confirmed acute fascioliasis, 9 patients with suspected fascioliasis, 27 patients with parasitic infections other than fascioliasis and 10 normal controls. Patients with fascioliasis were treated with TCZ as a single dose and followed up 1 and 4 months after therapy. The results showed that both IgM and IgG cystatin capture ELISAs had very high sensitivity and specificity of 100%. They were also able to diagnose early fascioliasis even before indirect haemagglutination test (IHAT) became positive. Within 4 months after treatment, only 20 patients (80%) with fascioliasis were cured as evidenced clinically, by normal eosinophil count and negative IHAT. After the first month of treatment, IgM and IgG cystatin capture ELISAs became negative in 16.6% and 83.3% of the cured cases respectively, reaching 95% for both assays after 4 months. They remained positive in 5 cases (20%) not responding to treatment.

Acute Disease↗
PubMed2001

[Activity of cathepsin B and cystatins in the placenta during EPH-gestosis].

Activity of cathepsin B using Bz-DL-Arg-pNA contents of SH-group by means of Ellman method, activity of cystatins against papain using casein as a substrate and contents of deoxyribonuclein acids by Burton method were determined in 64 placentas of pregnancies with EPH-gestosis and in 36 placentas of physiological pregnancies. The placentas from pregnancies with EPH-gestosis showed markedly higher activity of cathepsin B, no difference in the contents of SH-group, slightly higher activity of cystatins and they contain less deoxyribonucleic acids than the placentas from physiological pregnancies. The obtained results show that proteolytic--anti-proteolytic balance in placentas from pregnacies with EPH-gestosis is changed to the advantage of cathepsin B. This protease may in formation of structural and functional changes observed in placentas during EPH-gestosis.

Cathepsin B↗
PubMed2003

Cystatin capture-dot-enzyme-linked immunosorbent assay for immunodiagnosis and assessment of cure of experimental trichinellosis in mice.

Cystatin capture dot enzyme-linked immunosorbent assay (CC-dot ELISA) was evaluated as a new version of ELISA for diagnosis of trichinellosis by the detection of anti-Trichinella spiralis cysteine proteinase (CyP) IgG, using nitrocellulose membrane sensitized with cystatin as a capture reagent for CyP from T. spiralis muscle larvae excretory secretory products (ESP) without purification, compared to the detection of anti-T. spiralis IgG by conventional (conv.) ELISA, using whole ESP. Experimentally infected mice with light (100 larvae/mouse) and heavy (300 larvae/mouse) infections by T. spiralis larvae at 7, 14, 21 and 56 days post infection, and after flubendazole treatment were examined. As early as one week post infection CC-dot ELISA gave high positive rates of 86.6% and 100% in light and heavy infections, respectively, in contrast to negative results by conv. ELISA. CC-dot ELISA showed in light and heavy trichinellosis a higher efficiency in comparison to conv. ELISA (97.7% versus 67.7% and 98.8% versus 80%, respectively) and a higher overall sensitivity (96.6% versus 55% and 98.3% versus 73.3%, respectively). No cross reactions with sera of other parasitic infected or non infected control mice were recorded by CC-dot ELISA giving 100% specificity compared to 93.3% by conv. ELISA. After treatment, CC-dot ELISA gave positive results only in uncured mice with remaining muscle larvae, while conv. ELISA was positive in mice with and without remaining muscle larvae. CC-dot ELISA used lower quantities of antigen, was performed at room temperature, and read by naked eye in less than 2.5 hours.

Animals↗
PubMedDate not supplied

Chimeric enkephalin-cystatins: opioid binding and structure-activity relationships of inhibitory domains.

A chimeric inhibitor of cysteine proteinases, YGGFLQVVAGK.amide, was synthesized for use in examining SAR of its cystatin and opioid domains. Analogs were prepared by solid-phase syntheses and evaluated for inhibition of rat brain cathepsin L (E.C. 3.4.22.15) and papain (E.C.3.4.22.2), or binding to rat brain opioid receptors using tritiated DSLET (delta), DAGO (mu) and U69593 (kappa) in competitive binding assays. Cystatin consensus analogs QVVAGK- or N-Ac.QVVK.amide were weakly active as inhibitors, but were enhanced 10-fold to 20-fold with N-terminal L-, FL-, GFL-, GGFL-, or YGGFL, yielding Ki 3 microM and 26 microM for cathepsin L and papain, respectively. YGGFL itself was inactive, but expressed inhibition with N-terminal Q-, QV-, QVV-, etc. Highest activity was found with YGGFLQVK.amide (Ki, 2 microM). N-Dns analogs differentially increased inhibition for papain, whereas an N-Ac-cyclic peptide, N-Ac-CKYGGFLQVVKC.amide, was 2-fold to 10-fold less potent than the two-domain inhibitor. Chimeric peptides containing YGGFL were equipotent as delta-ligands (Ki, 1.7-3.7 nM), but were 2-fold to 10-fold more potent as mu ligands (3.7-11 nM) versus YGGFL itself (37 nM). Two analogs with -QVVAK- and -QVK.amide expressed kappa-binding properties. These data demonstrate the feasibility of using chimeric peptides as probes for exploring enzyme catalysis, and the potential for targeting inhibitors to endosomal or lysosomal compartments via receptor-mediated uptake in cells.

Amino Acid Sequence↗
PubMed2005

Serodiagnosis of human fascioliasis by a cystatin capture enzyme-linked immunosorbent assay with recombinant Fasciola gigantica cathepsin L antigen.

Cathepsin L1, a cysteine protease secreted by the gastrodermis of juvenile and adult Fasciola gigantica, was expressed in Escherichia coli as a calmodulin binding peptide fusion protein with a molecular mass of approximately 35 kD. The recombinant cathepsin L1 (rCTL1) was tested for its antigenic potential in a cystatin capture enzyme-linked immunosorbent assay (ELISA) to diagnose human fascioliasis. The ELISA plates were sensitized with chicken egg cystatin and incubated with bacterial lysates containing the recombinant protein before the standard ELISA procedures were performed. Analysis of the sera of 13 patients infected with F. gigantica (group 1), 204 patients with other parasitic infections (group 2), 32 cholangiocarcinoma patients (group 3), and 42 healthy controls (group 4) showed that the sensitivity, specificity, accuracy, positive predictive value, and negative predictive value of this ELISA using rCTL1 were 100%, 98.92%, 98.97%, 81.25%, and 100%, respectively. These results indicate that this assay has high sensitivity and specificity in the diagnosis of human fascioliasis. In addition, we have produced sufficient amounts of antigen for use in diagnosis.

Animals↗
PubMed1994

Urinary excretion of cathepsin B and cystatins as parameters of tubular damage.

The urinary excretion of the lysosomal hydrolases cathepsin B and beta-N-acetylglucosaminidase (beta-NAG) was compared with the tubular activities of these enzymes in remnant kidneys 16 weeks after subtotal nephrectomy (5/6 NX) or unilateral nephrectomy (UNX), as well as in kidneys from diabetic rats. In addition, the urinary excretion of the low-molecular weight protein cystatins, inhibitors of lysosomal cathepsins, was also followed in these animals. The urinary excretion of cathepsin B and beta-NAG was significantly enhanced in all three models of renal disease. The highest excretion rates for these enzymes were found in diabetic animals (cathepsin B: 4-fold; beta-NAG: more than a 10-fold increase over respective controls). In terms of tubular enzyme activities, tissue activities of both hydrolases were reduced in the remnant kidney after 5/6 NX, while in UNX and diabetes only cathepsin B activity was decreased. The urinary excretion of cystatins was enhanced in all three animal models, particularly in 5/6 nephrectomized rats, where a 40-fold increment over control animals was observed. Taken together, these findings indicate that there was severe tubular damage in the remnant kidney after 5/6 NX (reduced tubular enzyme activities, enzymuria and severely compromised tubular protein reabsorption). Furthermore, considerable enzymuria and disturbed protein reabsorption in early diabetes suggest tubular dysfunction before signs of glomerular damage become evident.

Acetylglucosaminidase↗
PubMed1998

[The role of cystatin in neoplastic progression and diagnosis].

Possible role of the cystatins, natural and specific inhibitors of the cysteine peptidases in cancer invasion is described. The value of cystatins determinations in serum and tissue extracts as diagnostic and prognostic parameters in human cancers is also presented.

Animals↗