Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P01034 (cystatin C)
 3,397 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cystatins are a superfamily of low Ki cysteine proteinase inhibitors found in both plants and animals. Cystatin C, a secreted molecule of this family, is of interest from biochemical and evolutionary points of view, and also has biotechnological applications. Recently we cloned and sequenced the cDNA for rainbow trout (Oncorhynchus mykiss) cystatin C [Li et al., 1998. Molecular cloning, sequence analysis and expression distribution of rainbow trout (Oncorhynchus mykiss) cystatin C. Comp. Biochem. Phys. B 121, 135-143]. To explore the relationship between protein structure and function of trout cystatin C, we established a bacterial system for expression of the protein. Trout cystatin C expressed in the cytoplasm of bacterial cells did not have detectable protease inhibitory activity. Activity was regained by Ni-NTA chromatography under denaturing conditions followed by dialysis-based refolding. Titration of purified cystatin C preparations with papain indicated that approximately 20% of the total protein had been converted to the active form after one refolding cycle. Expression levels were 3-5 mg/l. The protease-inhibitory properties of recombinant trout cystatin C were similar to those of human and chicken cystatin C derived from biological sources and recombinant cystatin C derived from rat and mouse genes. The Ki for papain was 1.2 x 10(-15) M, exhibiting the high affinity binding unique to this family of protease inhibitors.
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PMID:Rainbow trout (Oncorhynchus mykiss) cystatin C: expression in Escherichia coli and properties of the recombinant protease inhibitor. 1090 62

The parasitic protozoan Trypanosoma cruzi employs multiple molecular strategies to invade a broad range of nonphagocytic cells. Here we demonstrate that the invasion of human primary umbilical vein endothelial cells (HUVECs) or Chinese hamster ovary (CHO) cells overexpressing the B(2) type of bradykinin receptor (CHO-B(2)R) by tissue culture trypomastigotes is subtly modulated by the combined activities of kininogens, kininogenases, and kinin-degrading peptidases. The presence of captopril, an inhibitor of bradykinin degradation by kininase II, drastically potentiated parasitic invasion of HUVECs and CHO-B(2)R, but not of mock-transfected CHO cells, whereas the B(2)R antagonist HOE 140 or monoclonal antibody MBK3 to bradykinin blocked these effects. Invasion competence correlated with the parasites' ability to liberate the short-lived kinins from cell-bound kininogen and to elicit vigorous intracellular free calcium ([Ca(2+)](i)) transients through B(2)R. Invasion was impaired by membrane-permeable cysteine proteinase inhibitors such as Z-(SBz)Cys-Phe-CHN(2) but not by the hydrophilic inhibitor 1-trans-epoxysuccinyl-l-leucyl-amido-(4-guanidino) butane or cystatin C, suggesting that kinin release is confined to secluded spaces formed by juxtaposition of host cell and parasite plasma membranes. Analysis of trypomastigote transfectants expressing various cysteine proteinase isoforms showed that invasion competence is linked to the kinin releasing activity of cruzipain, herein proposed as a factor of virulence in Chagas' disease.
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PMID:Host cell invasion by Trypanosoma cruzi is potentiated by activation of bradykinin B(2) receptors. 1106 78

Variant human cystatin C (L68Q) is an amyloidogenic protein. It deposits in the cerebral vasculature of Icelandic patients with cerebral amyloid angiopathy, leading to stroke. Wild-type and variant cystatin C are cysteine proteinase inhibitors which form concentration dependent inactive dimers; however, variant cystatin C dimerizes at lower concentrations and has an increased susceptibility to a serine protease. We studied the effect of the L68Q amino acid substitution on cystatin C properties, utilizing full length cystatin C purified in mild conditions from media of cells stably transfected with either the wild-type or variant cystatin C genes. The variant cystatin C forms fibrils in vitro detectable by electron microscopy in conditions in which the wild-type protein forms amorphous aggregates. We also show by circular dichroism, steady-state fluorescence and Fourier-transformed infrared spectroscopy that the amino acid substitution modifies cystatin C structure by destabilizing alpha-helical structures and exposing the tryptophan residue to a more polar environment, yielding a more unfolded molecule. These spectral changes demonstrate that variant cystatin C has a three-dimensional structure different from that of the wild-type protein. The structural differences between variant and wild-type cystatin C account for the susceptibility of the variant protein to unfolding, proteolysis and fibrillogenesis.
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PMID:Distinct properties of wild-type and the amyloidogenic human cystatin C variant of hereditary cerebral hemorrhage with amyloidosis, Icelandic type. 1129 25

A sandwich-type ELISA has been developed for quantification of the complex between the cysteine proteinase cathepsin B (CB) and its reversible tight-binding inhibitor cystatin C (CC) in normal and pathological sera. The assay is based on a combination of catching Ab (3E1), raised against CB, and a horseradish peroxidase-labelled detection Ab (1A2), raised against CC. Only the CB/CC complex is able to evoke a signal in this assay. The detection limit of the assay was 15.5 nM and the working range between 31.3-200 nM. The within and between-run coefficients of variance (CV) varied from 4.7% to 9.4% and 11% to 12.8%, respectively, demonstrating satisfactory reproducibility of the method. The concentration of the CB/CC complex was determined in sera from 90 healthy controls, 32 patients with non-cancerous lung diseases, 148 patients with lung and 32 patients with colorectal cancer. The CB/CC complex was significantly less abundant in sera of patients bearing malignant lung tumours than in those with non-cancerous lung diseases or healthy controls (p<0.001). In colorectal cancer sera its level was significantly lower in advanced stages C and D than in early Dukes' stages A and B (p=0.02). Our results show that the increased levels of CB in malignant sera are not impaired effectively by CC and support the hypothesis of hindered inhibitory capability during cancer progression.
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PMID:Cathepsin B/cystatin C complex levels in sera from patients with lung and colorectal cancer. 1151 34

The activities'of the lysosomal cysteine proteinases cathepsin B and L are regulated by their endogenous inhibitors, stefins A and B, and cystatin C, and their imbalance may be associated with increased invasiveness and development of the malignant cell phenotype. The aim of this study was to investigate mRNA, protein and activity levels of the above proteins in relation to in vitro invasiveness and to the reported in vivo tumorigenicity of four human breast tumor cell lines: the spontaneously immortalized cell line MCF10A, its c-Ha-ras transfectant MCF10AT, and two tumorigenic derivative cell lines, MCF10AT-Ca1a and MCF10AT-Ca1d. Invasiveness did not correlate with tumorigenicity, since the MCF10AT cell was the most invasive and the remaining three were at about half of its level. Cathepsin B expression paralleled the in vitro invasiveness through matrigel at all levels of expression, but cathepsin L did not. Stefin levels were elevated several-fold in the tumorigenic cell lines, but not in MCF10AT. The hypothesis that cathepsin B plays an active role in the invasion of breast cancer cell lines was confirmed by the fact that synthetic cysteine proteinase inhibitors, particularly those selective for cathepsin B, significantly reduced the invasion of the MCF10AT cells.
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PMID:Invasiveness of transformed human breast epithelial cell lines is related to cathepsin B and inhibited by cysteine proteinase inhibitors. 1271 95

Replacement of the three N-terminal residues preceding the conserved Gly of cystatin A by the corresponding 10-residue long segment of cystatin C increased the affinity of the inhibitor for the major lysosomal cysteine proteinase, cathepsin B, by approximately 15-fold. This tighter binding was predominantly due to a higher overall association rate constant. Characterization of the interaction with an inactive Cys29 to Ala variant of cathepsin B indicated that the higher rate constant was a result of an increased ability of the N-terminal region of the chimeric inhibitor to promote displacement of the cathepsin B occluding loop in the second binding step. The low dissociation rate constant for the binding of cystatin A to cathepsin B was retained by the chimeric inhibitor, which therefore had a higher affinity for this enzyme than any natural cystatin identified so far. In contrast, the N-terminal substitution negligibly affected the ability of cystatin A to inhibit papain. However, substitutions of Gly75 in the second binding loop of cystatin A by Trp or His, making the loop similar to those of cystatins C or B, respectively, increased the affinity for papain by approximately 10-fold. This enhanced affinity was due to both a higher association rate constant and a lower dissociation rate constant. Modeling of complexes between the two variants and papain indicated the possibility of favorable interactions being established between the substituting residues and the enzyme. The second-loop substitutions negligibly affected or moderately reduced the affinity for cathepsin B. Together, these results show that the inhibitory ability of cystatins can be substantially improved by protein engineering.
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PMID:Grafting of features of cystatins C or B into the N-terminal region or second binding loop of cystatin A (stefin A) substantially enhances inhibition of cysteine proteinases. 1450 83

Cystatin C is a secreted inhibitor of cysteine proteinases that participates in extracellular matrix remodeling. Whether hormones affect its expression in the vagina was unknown. Consequently, we examined the effects of estradiol (E(2)), progesterone (P), and raloxifene on vaginal cystatin C in rhesus macaques. In experiment 1, ovariectomized animals were treated sequentially with E(2) (14 d) and E(2) + P (14 d) to induce 28-d menstrual cycles. Vaginal samples were collected on d 6, 8, 14, and 28 of the induced cycle. Some cycled animals were deprived of both E(2) + P for 28 d. In experiment 2, ovariectomized animals were treated for 5 months with E(2) alone, E(2) + P, raloxifene, or left untreated. Total RNA from the vaginal wall was analyzed for the cystatin C transcript with a commercially prepared cDNA array and semiquantitative RT-PCR. Vaginal cryosections were analyzed by in situ hybridization for cystatin C transcript and by immunocytochemistry for the protein. E(2) treatment significantly (5-fold; P < 0.05) increased expression of cystatin C transcript over the levels in the hormone-deprived controls, and cotreatment with P (E(2) + P) blocked this effect. Raloxifene treatment did not affect cystatin C expression. In situ hybridization and immunocytochemistry revealed that cystatin C was localized in fibroblasts and smooth muscle cells throughout the vaginal wall but not in smooth muscle cells of arteries or levator ani myocytes. In summary, E(2) increased vaginal cystatin C expression in the fibroblasts and smooth muscle bundles, P suppressed this effect, and raloxifene had no effects on cystatin C. Elevated cystatin C, by suppressing cysteine proteinase activity, may strengthen the vaginal wall and mitigate the potential for pelvic floor prolapse.
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PMID:Estrogen enhances cystatin C expression in the macaque vagina. 1476 9

The effects of cystatin C and other cysteine proteinase inhibitors on osteoclast formation and differentiation have been investigated. Cystatin C decreased osteoclast formation stimulated by parathyroid hormone (PTH), 1,25(OH)2-vitamin D3 or interleukin-6 (IL-6) (in the presence of its soluble receptor) as assessed by the number of tartrate-resistant acid phosphatase (TRAP+) multinucleated cells in mouse bone marrow cultures. The inhibitory effect was associated with decreased mRNA expression for the calcitonin receptor as well as decreased number of specific binding sites for 125I-calcitonin, and without any effect on the mRNA expression of receptor activator of nuclear factor kappaB (NF-kappaB) ligand (RANKL). Similarly, the cysteine proteinase inhibitors leupeptin, E-64 and benzyloxycarbonyl-Phe-Ala-diazomethane (Z-FA-CHN2) decreased PTH-stimulated formation of TRAP+ multinucleated cells and binding of 125I-calcitonin. A peptidyl derivative synthesized to mimic part of the proteinase-binding site of cystatin C (benzyloxycarbonyl-Arg-Leu-Val-Gly-diazomethane, or Z-RLVG-CHN2) also decreased PTH-stimulated osteoclast formation. In a 9-day culture, addition of cystatin C during the last 5 days was sufficient to cause substantial inhibition of osteoclast formation. Cystatin C-induced decrease of osteoclast formation was associated with enhanced number of F4/80-positive macrophages and increased mRNA expression of the macrophage receptor c-fms in the bone marrow culture. Osteoclast formation in mouse bone marrow cultures as well as in mouse spleen cell cultures, stimulated by macrophage colony-stimulating factor (M-CSF) and RANKL was also decreased by different cysteine proteinase inhibitors. In addition, cystatin C inhibited M-CSF/RANKL induction of calcitonin receptor mRNA in spleen cell cultures. The inhibitory effect by cystatin C in spleen cells was associated with decreased mRNA expression of RANK and the transcription factor NFAT2. It is concluded that cysteine proteinase inhibitors decrease formation of osteoclasts by interfering at a late stage of pre-osteoclast differentiation.
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PMID:Osteoclastogenesis is decreased by cysteine proteinase inhibitors. 1500 89

To determine the role of the cysteine proteinase inhibitor cystatin C in the invasive behavior of squamous cell carcinoma of the head and neck (SCCHN), Cystatin C protein level was measured in 82 pairs of primary tumour tissue and adjacent noncancerous mucosa, using the enzyme-linked immunosorbent assay. The median level of cystatin C in tumour tissue was 1.18 times lower than that in corresponding mucosa (P=0.031). In normal mucosa samples, the cystatin C level was influenced by the site of sampling: it was lower in nonlaryngeal tissue samples (oral cavity, oro- or hypopharynx) than in laryngeal samples (P=0.004). The tumour cystatin C level correlated inversely with pN-stage (P=0.047), whereas a trend of lower cystatin C levels was observed in the group with extranodal tumour extension compared to those with no extranodal spread (P=0.069). In univariate analysis, the patients with low tumour cystatin C levels exhibited poor disease-free survival (DFS, P=0.013) and disease-specific survival (DSS, P=0.013). In multivariate analysis, the most powerful predictor of survival was pN-stage (DFS: P=0.040, HR 2.78; DSS: P=0.011, HR 4.36,), followed by the cystatin C level (DFS: P=0.043, HR 0.22; DSS: P=0.067, HR 0.25). When comparing the prognostic strength of cystatin C to that of stefin A, another cysteine proteinase inhibitor, which emerged as the most significant prognosticator for survival in our previous study analysing the same cohort of patients, stefin A proved to be significantly more reliable predictor for both DFS and DSS than cystatin C. Our results indicate that cystatin C is implicated in the invasive behavior of SCCHN, and that there are variations in regulation of proteolytic pathways under nonmalignant conditions, inherent to individual subsites inside the upper aerodigestive tract. The correlation between high cystatin C levels and improved survival concurs with the concept of the protective role of high levels of cysteine proteinase inhibitors in tissue homogenates that has been previously suggested by the survival results in breast and lung carcinoma as well as SCCHN.
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PMID:Cysteine proteinase inhibitor cystatin C in squamous cell carcinoma of the head and neck: relation to prognosis. 1513 78

Development of murine HA-1 hepatoma was accompanied by increased activity of cathepsin B (in ascitic cells), cathepsin D (in ascitic fluid) and increased activity of procathepsin B. There were some changes of cysteine proteinases in liver and spleen, not involved directly into tumor growth. The most prominent changes included the decreased level of cysteine proteinase inhibitors cystatin C and stefin A in ascitic cells (and to a lesser degree in liver tissue). During tumor development serum cystatin C concentration decreased by 3-times compared to intact mice. Treatment by antitumor drug Ukraine increased life span of mice with HA-1 hepatoma (transplanted intravenously), decreased the increment of tumor weight. In ascite such treatment caused a decrease of number of tumor cells and an increase of number of macrophages. Ukraie (administered once or 5-times in a dose of 0.5 mg per mice) increased cystatin C level, revealing protective mechanism of action.
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PMID:[Cysteine proteinases and their inhibitors in the development of mouse HA-1 hepatoma and antineoplastic therapy]. 1517 24


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