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.

SourceResearch connectionFrom yearThrough year

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

At least 1,027 records · Page 57Linked to original sources

PubMed1993

A sequence variation in the human cystatin D gene resulting in an amino acid (Cys/Arg) polymorphism at the protein level.

A polymorphism in the coding region of the human cystatin D gene has been detected by direct sequencing of amplified DNA from different individuals. The variation, resulting from a T/C transition in exon 1 of the gene, causes an amino acid variation, Cys/Arg, at the protein level. An allele-specific oligonucleotide hybridization assay was developed and used to demonstrate this polymorphism in the population. the deduced frequencies were 0.55 and 0.45 for the Cys and Arg variant-encoding alleles, respectively.

Amino Acid Sequence↗

PubMed1996

The possible place of cathepsins and cystatins in the puzzle of Alzheimer disease: a review.

Lysosomal proteinases (cathepsins) and their endogenous inhibitors (cystatins) have been found to be closely associated with senile plaques, cerebrovascular amyloid deposits, and neurofibrillary tangles in Alzheimer disease (AD). Further, profound changes in the lysosomal system seem to be an early event in "at-risk" neurons of AD brains. There is an ongoing controversy as to whether lysosome-associated proteolytic mechanisms are causally related to the development and/or further progression of the disease. The present article deals with some arguments "pro" and "contra" an involvement of the endosomal/lysosomal pathway in amyloidogenesis as a cardinal process in AD. Other putative targets of acidic proteinases and their natural inhibitors in the pathogenesis of AD (such as formation of neurofibrillary tangles and regulation of apolipoprotein E) are also discussed.

Alzheimer Disease↗

PubMed1998

Binding of the pili of Pseudomonas aeruginosa to a low-molecular-weight mucin and neutral cystatin of human submandibular-sublingual saliva.

In previous experiments, we have shown that [125I]pili of Pseudomonas aeruginosa exhibited specific binding to a low-molecular-weight mucin (MG2) of human submandibular-sublingual saliva (HSMSL; Reddy MS, Levine MJ, Paranchych W. Crit Rev Oral Biol Med 4:315-323, 1993). In the present study, I have utilized unlabeled pili and immunostaining to identify the receptor molecules in HSMSL. In addition to MG2, pili also bound to neutral cystatin (CsnSN). Binding of unlabeled pili to MG2 and CsnSN could be abolished by treatment of HSMSL with trypsin to hydrolyze the peptide moieties or N-acetylation to neutralize the positive charges of the lysine residues. Reductive methylation of HSMSL, which modifies the lysine residues to methyl lysines while retaining the positive charges, did not affect the binding of pili to either MG2 or CsnSN. Further, pili also exhibited binding to a recombinant MG2 peptide (aa 1-86). Collectively, the data suggested that a protein-to-protein interaction via electrostatic forces mediates the binding of the pili to MG2 and CsnSN. Iodination of pili, which modifies tyrosine-24 and/or -27 residues to O-iodotyrosine(s), abolished its binding to CsnSN but not to MG2. These results suggested that the conformation of pili also plays a role in interaction with CsnSN. Conformational change(s) of pili induced by iodination also made it susceptible to hydrolysis with trypsin.

Cystatins↗

PubMed1998

Expression of cystatin-related protein and of the C3-component of prostatic-binding protein during postnatal development in the rat ventral prostate and lacrimal gland.

The expression of cystatin-related protein (CRP) and of the C3-component of prostatic-binding protein (PBP) during postnatal development of the rat was studied by Northern blotting, dot blot and in situ hybridisation, and by radioimmunoassay or immunoblotting. In intact male rats, very little or no PBP-C3 could be detected in the prostate at 10 days, but at 20 days there was already strong expression. By in situ hybridisation, the first expression of C3 mRNA was observed at 13 days in the prostate and at 22 days in the lacrimal gland. For CRP, this occurred at 16 and 22 days, respectively. Neither CRP nor C3 was expressed in prepubertal male rats castrated at day 1 or day 10 or in female rats. Androgen treatment of intact male animals did not advance the expression of both mRNAs in the prostate, but did so in the lacrimal gland with first expression of C3 at 19 instead of 22 days and of CRP at 13 instead of 22 days. Identical values were obtained in female rats. Androgen treatment of castrated adult male rats resulted in a more rapid and homogeneous secondary induction. Positive immunostaining for the androgen receptor (AR) was observed in the lacrimal gland at 7 days, but its concentration, estimated by immunoblotting, was still low at 10 days. Maximal levels, reached at 30 days, were markedly higher in male than in female rats. In conclusion, CRP and C3 are induced by androgens in prepubertal rats. The time point of induction, however, is probably determined by other tissue and differentiation-dependent factors in addition to androgens and the AR.

Aging↗

PubMed2005

Comparison of guanidine hydrochloride (GdnHCl) and urea denaturation on inactivation and unfolding of human placental cystatin (HPC).

The activity and conformational change of human placental cystatin (HPC), a low molecular weight thiol proteinase inhibitor (12,500) has been investigated in presence of guanidine hydrochloride (GdnHCl) and urea. The denaturation of HPC was followed by activity measurements, fluorescence spectroscopy and Circular Dichroism (CD) studies. Increasing the denaturant concentration significantly enhanced the inactivation and unfolding of HPC. The enzyme was 50% inactivated at 1.5 M GdnHCl or 3 M urea. Up to 1.5 M GdnHCl concentration there was quenching of fluorescence intensity compared to native form however at 2 M concentration intensity increased and emission maxima had 5 nm red shift with complete unfolding in 4-6 M range. The mid point of transition was in the region of 1.5-2 M. In case of urea denaturation, the fluorescence intensity increased gradually with increase in the concentration of denaturant. The protein unfolded completely in 6-8 M concentration of urea with a mid-point of transition at 3 M. CD spectroscopy shows that the ellipticity of HPC has increased compared to that of native up to 1.5 M GdnHCl and then there is gradual decrease in ellipticity from 2 to 5 M concentration. At 6 M GdnHCl the protein had random coil conformation. For urea the ellipticity decreases with increase in concentration showing a sigmoidal shaped transition curve with little change up to 1 M urea. The protein greatly loses its structure at 6 M urea and at 8 M it is a random coil. The urea induced denaturation follows two-state rule in which Native-->Denatured state transition occurs in a single step whereas in case of GdnHCl, intermediates or non-native states are observed at lower concentrations of denaturant. These intermediate states are possibly due to stabilizing properties of guanidine cation (Gdn+) at lower concentrations, whereas at higher concentrations it acts as a classical denaturant.

Cystatins↗

PubMed1990

Simulation of the inhibitory cystatin surface by a synthetic peptide.

An inhibitory dodecameric peptide was designed which tentatively mimics the inhibitory site of cystatin C-like structures. Succinylated and mansylated derivatives were also synthesised and assayed for their inhibiting properties towards papain and rat cathepsins B, H and L. All peptides preferentially inhibit cathepsin L and papain as their naturally occurring inhibitor model. A significant increase in inhibition was obtained after mansylation of the crude peptide with Ki values in the micromolar or 0.1 micromolar range. The use and interest of such peptide inhibitors are discussed.

Amino Acid Sequence↗

PubMed1989

Stem bromelain: amino acid sequence and implications for weak binding of cystatin.

The amino acid sequence of stem bromelain, the major cysteine proteinase from pineapple stem is described. It shows that the enzyme is a member of the papain superfamily of cysteine proteinases, but is not very closely related to any other known member of this group. The sequence shows mutation or deletion of several residues that have been conserved in cysteine proteinases examined previously, including Asn-175 (papain). We suggest that some of these changes have the effect of altering the active-site geometry of stem bromelain, and that this accounts for the resistance of the enzyme to inhibition by cystatins and E-64[L-3-carboxy-2,3-trans-epoxypropionylleucylamido(4-guanidino)b utane].

Amino Acid Sequence↗

PubMed1992

Biphasic transition curve on denaturation of chicken cystatin by guanidinium chloride. Evidence for an independently unfolding structural region.

Far-ultraviolet circular dichroism and tryptophan fluorescence measurements showed that the reversible unfolding of the cysteine proteinase inhibitor, chicken cystatin, by guanidinium chloride is a two-step process with transition midpoints at approximately 3.4 and approximately 5.4 M denaturant. The partially unfolded intermediate had both far- and near-ultraviolet circular dichroism and fluorescence emission spectra comparable to those of the native protein. The largely retained tertiary structure suggests that the intermediate represents a species in which a separate region of lower stability has been unfolded, rather than an intermediate of the 'molten globule' type. Such a structurally independent region is apparent in the three-dimensional structure of the inhibitor.

Animals↗

PubMed1995

The removal of 2-oxoacyl residues from the N-terminus of peptides and cystatin in non-denaturing conditions.

The N-terminal residue of a protein or peptide may be converted into a 2-oxoacyl group by non-enzymic transamination. This group may then be removed, to obtain the peptide chain shortened by one residue, by treatment with phenylene-1,2-diamine. Hitherto this scission has required a pH of 4-5, but we find that the reaction will proceed well at pH 7 in the presence of concentrated phosphate buffer. We describe a method using reverse-phase HPLC for determining the extent of scission in model peptides; this method also allows products to be isolated and identified. The new scission conditions have been tested by removing the N-terminal residue from cystatin, an inhibitor of cysteine peptidases; electrospray mass spectrometry was used to assess how this protein reacted.

Amino Acid Sequence↗

PubMed1994

New substrates of papain, based on the conserved sequence of natural inhibitors of the cystatin family.

A series of peptide substrates with different fluorogenic leaving groups has been synthesized. The peptide moiety in these substrates mimics a highly conserved sequence (QVVAG) in the natural reversible inhibitors of cysteine proteinases, the cystatins, that participates to the tight binding of target proteinases. This sequence is invariably cleaved at the A-G bond when synthetic peptides containing it were incubated with papain. AEC and AMC fluorophores were therefore attached to the Ala residue to construct new substrates for cysteine proteinases. The solubility of the resulting substrates was improved by attaching a N-terminal gluconoyl group, or by introducing an arginyl residue at P5 (nomenclature of Schechter I, Berger A (1967) Biochem Biophys Res Commun 27, 157-162). Neither induced significant changes in the kcat/Km values with papain. Those values were all in the 10(5) M-1 s-1 range. The kcat/Km was increased 10-50-fold by using substrates with intramolecularly quenched fluorescence. With these, the enzyme specificity on both sides of the scissile bond can be investigated. The substrate Abz-QVVAGA-EDDnp is among the most sensitive papain substrates ever reported, with a kcat/Km value of 29 10(6) M-1 s-1. The positioning and conformation of the bound QVVA moiety within the active site of papain were predicted by molecular modelling using the X-ray coordinates of a peptide inhibitor-papain complex.

Amino Acid Sequence↗

PubMed1996

Androgenic induction of cystatin-related protein and the C3 component of prostatic binding protein in primary cultures from the rat lacrimal gland.

Cystatin-related protein (CRP) and the C3 component of prostatic binding protein (C3) are synthesized in vivo under androgen control in the lacrimal gland and ventral prostate of adult male rats [1,2]. Androgen administration to female or 7-day castrated male rats, which do not express CRP, can induce its synthesis [3]. In this study, we show androgen-dependent expression of CRP1 and C3 in primary cultures of acinar cells of the lacrimal gland of female rats. Addition of androgens to the culture medium results in the synthesis and secretion of both proteins in a time- and dose-dependent way. Estradiol or progesterone are unable to induce their expression. Dexamethasone in low concentrations and present as a basal component of the serum free defined medium, is needed to sensitize the culture system for androgens. In high concentrations, this synthetic glucocorticoid seems to play a similar role as androgens in CRP1 and C3 regulation.

Androgen-Binding Protein↗

PubMed1996

Purification and molecular cloning of carp ovarian cystatin.

The ovarian fluid of carp consists of many components. Using the antiserum against carp serum, Western blot analysis of ovarian fluid was done in order to distinguish substances synthesized by the ovary from those derived from the serum. Several ovary-specific substances were detected including a protein of 12 kDa (p12), which was purified to homogeneity. Purified p12 displays a single band in SDS-PAGE under nonreducing condition and it can inhibit the enzymatic activity of papain with an apparent inhibition constant of 0.01 nM. The primary structure of p12 was partially determined by Edman degradation and fully elucidated by molecular cloning. A cDNA of 531 bp encoding p12 was obtained. The precursor of p12 has 129 residues, including a signal peptide of 18 residues and a mature protein of 111 residues. The N- and C-terminus of p12 are threonine and methionine, respectively. The p12 shares many common features of the family 2 cystatins of other species, including the similarity of the protein size (in the range of 110 to 120 residues), the presence of 4 cysteine residues and the occurrence of invariant residues throughout the molecule.

Amino Acid Sequence↗

PubMed1993

Isolation and characterization of the chicken cystatin-encoding gene: mapping transcription start point and polyadenylation sites.

A 6.9-kb fragment containing coding sequences for chicken egg white cystatin (CsnEW) was isolated from a chicken genomic library using the CsnEW cDNA as a probe. The gene is approximately 2.4 kb in length; it contains three exons, two introns and two polyadenylation signals. The exon-intron arrangement corresponds exactly with those of other members of the Csn superfamily. The sequence of the 5' flanking region contains two SP1-binding sites and a high G+C content suggestive of a housekeeping gene. All tissues studied express CsnEW mRNA; Northern analysis showed that CsnEW mRNA levels are most abundant in lung and least abundant in liver and spleen. Mapping of the 3' end of the CsnEW mRNA isolated from brain tissue resolved two CsnEW mRNA species. The larger transcript resulting from the use of the second polyadenylation signal was more abundant than the smaller transcript. Determination of the transcription start point (tsp) of CsnEW mRNA by primer extension and RNase protection assays showed that CsnEW mRNA from a number of chicken tissues was approximately 40-50 nucleotides shorter than that predicted from the CsnEW cDNA isolated from chicken oviduct.

Amino Acid Sequence↗

PubMed1993

Sequence of the proteinase-inhibitor cystatin homologue from the pollen of Ambrosia artemisiifolia (short ragweed).

The nucleotide sequence of a cDNA (designated IPC1/5) encoding a cystatin (Cyt) proteinase-inhibitor homologue from short ragweed (Ambrosia artemisiifolia) pollen was determined and compared to other plant and animal Cyt. The absence of disulfide bonds in the predicted translation product of the IPC1/5 sequence suggests that it most resembles family-I members of the Cyt superfamily. Significant amino acid (aa) sequence identity was found when comparing the translated sequence of IPC1/5 to rice seed Cyt, human keratocyte Cyt A and human liver Cyt B.

Allergens↗

PubMed1995

Inhibition of cysteine proteinases by Carica papaya cystatin produced in Escherichia coli.

A papaya cystatin (Cst)-encoding cDNA clone was isolated from a papaya leaf cDNA library and the active protein produced in Escherichia coli. The amino-acid sequence reveals a protein of 11,262 Da with over 40% identity to other published plant Cst. Unique features of the papaya Cst include a single Cys residue, variation in the papain-binding region, and the first reported inhibition of papaya proteinase IV by a Cst.

Base Sequence↗

PubMed1994

Androgen-dependent expression of cystatin-related protein (CRP) in the exorbital lacrimal gland of the rat.

Cystatin-related protein (CRP), also known as 20 (22)-kDa glycoprotein is expressed not only in the ventral prostate, but also in the lacrimal gland of adult male rats. In this study the expression of CRP in androgen-treated female animals is studied. CRP mRNA is absent in the lacrimal gland of untreated adult female rats, but can be induced by androgens, although this induction is slower than in castrated male rats. Estradiol, progesterone or glucocorticoids have no effect. In testicular feminized rats, however, CRP mRNA is not induced in the lacrimal gland by androgens. At the protein level, the presence of CRP in tears of adult male rats is demonstrated. In female animals or castrated male animals CRP can be induced by androgens in a dose-dependent way. Here also the induction is slower in female rats, even during secondary induction after previous full stimulation by androgens. These results indicate that androgens and a functionally normal androgen receptor are essential for the expression of CRP in the lacrimal gland. The time course of induction depends on the dose of androgens, the previous contact with androgens, the duration of the androgen-free interval and the sex of the animals.

Androgen-Insensitivity Syndrome↗

PubMed2004

The N-terminus moiety of the cystatin SmCys from Schistosoma mansoni regulates its inhibitory activity in vitro and in vivo.

The complete sequence of SmCys, a cystatin expressed by Schistosoma mansoni, was obtained. Constructs of SmCys consisting of deletions of 10 and 20 amino acid residues from the N-terminal of the full length recombinant protein, were cloned in the pQE-30 vector, expressed in Escherichia coli and assayed for inhibitory activity against papain. Kinetic analysis showed that SmCys -10 and SmCys -20 had K(i) values of 0.7391 and 4.9154, respectively, as compared to 0.0647, displayed by the full length recombinant. Protease inhibition by SmCys was also observed in vivo. When the recombinant products were incubated during 7 days with live schistosomula in the presence of red blood cells, only the full length product could completely inhibit the formation of haemozoin, a dark pigment formed as a by-product of haemoglobin digestion. The sequence data of the recombinant SmCys proteins were used for the construction of molecular models, which were then subjected to molecular dynamics for 2ns. In comparison to the full length, the models corresponding to the truncated constructs, showed a distinctive change on the surface charge distribution. This parameter was more pronounced in SmCys -20, which also displayed a significant displacement of the inhibitory domain, a result which could explain the kinetic data in terms of the loss of attachment sites. These changes correlated well with the progressive lack of inhibition observed for the recombinant deletion constructs, in vitro and in vivo.

Amino Acid Sequence↗

PubMed2005

An inflammatory role for the mammalian carboxypeptidase inhibitor latexin: relationship to cystatins and the tumor suppressor TIG1.

Latexin, the only known mammalian carboxypeptidase inhibitor, has no detectable sequence similarity with plant and parasite inhibitors, but it is related to a human putative tumor suppressor protein, TIG1. Latexin is expressed in the developing brain, and we find that it plays a role in inflammation, as it is expressed at high levels and is inducible in macrophages in concert with other protease inhibitors and potential protease targets. The crystal structure of mouse latexin, solved at 1.83 A resolution, shows no structural relationship with other carboxypeptidase inhibitors. Furthermore, despite a lack of detectable sequence duplication, the structure incorporates two topologically analogous domains related by pseudo two-fold symmetry. Surprisingly, these domains share a cystatin fold architecture found in proteins that inhibit cysteine proteases, suggesting an evolutionary and possibly functional relationship. The structure of the tumor suppressor protein TIG1 was modeled, revealing its putative membrane binding surface.

Amino Acid Motifs↗