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)

The peptidyl diazomethanes Cbz-Gly-CHN2, Boc-Val-Gly-CHN2, H-Leu-Val-Gly-CHN2, Cbz-Leu-Val-Gly-CHN2 and Cbz-Arg-Leu-Val-Gly-CHN2, with peptidyl portions modelled after the proposed cysteine proteinase interacting N-terminal segment of human cystatin C, were synthesized. Their efficiency as cysteine proteinase inhibitors was tested against papain, human cathepsin B and bovine cathepsin B. All, except Cbz-Gly-CHN2, were found to be irreversible inhibitors of the tested enzymes. Each addition of an amino acid residue to their peptidyl portions resulted in an increased inhibition rate of all three enzymes. These data suggest that the arginyl residue of the tetrapeptidyl diazomethane, and also the corresponding arginyl residue in native cystatin C, interact with a S4 substrate pocket subsite of both papain and cathepsin B. The most efficient inhibitor, Cbz-Arg-Leu-Val-Gly-CHN2, inhibited papain and cathepsin B with rate constants of the same order of magnitude as those for L-3-carboxy-trans-2,3-epoxypropionyl-leucylamido-(4-guanidin o)butane (E-64). The high water-solubility of Cbz-Arg-Leu-Val-Gly-CHN2 allowing it to be dissolved to molar concentrations without use of non-physiological additives, makes it suitable for in vitro and in vivo cysteine proteinase inhibition studies.
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PMID:Cystatin C based peptidyl diazomethanes as cysteine proteinase inhibitors: influence of the peptidyl chain length. 128 27

Molecular analysis of the human beta-galactosidase gene revealed six different mutations in 10 of 11 Japanese GM1-gangliosidosis patients. They were the only abnormalities in each allele examined in this study. A 165-nucleotide duplication (positions 1103-1267) was found in two infantile patients, producing an abnormally large mRNA; one patient was probably a homozygote, and the other was a heterozygote of this mutation. The other two infantile patients had different mutations; a 123 Gly(GGG)----Arg(AGG) mutation in one patient and a 316 Tyr(TAT)----Cys(TGT) mutation in the other. A 201 Arg(CGC)----Cys(TGC) mutation, eliminating a BspMI site, was detected in a late-infantile/juvenile patient; the restriction-site analysis of amplified genomic DNA confirmed his heterozygosity for this mutation. A 51 Ile(ATC)----Thr(ACC) mutation was found in all five adult/chronic patients examined in this study. It created a SauI site, and restriction-site analysis confirmed that four patients were homozygous mutants. The other was a compound heterozygote for this mutation and another 457 Arg(CGA)----Gln(CAA) mutation. These mutant genes expressed markedly decreased or completely deficient enzyme activities in beta-galactosidase-deficient human fibroblasts transformed by adenovirus-SV40 recombinants. We conclude that gene mutations are heterogeneous in GM1-gangliosidosis but that the 51 Ile(ATC)----Thr(ACC) mutation is common among the Japanese adult/chronic cases. Genotype-phenotype correlations in GM1-gangliosidosis are briefly discussed.
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PMID:Human beta-galactosidase gene mutations in GM1-gangliosidosis: a common mutation among Japanese adult/chronic cases. 190

Leucocyte elastase in catalytic amounts was observed to rapidly cleave the Val-10-Gly-11 bond of the human cysteine-proteinase inhibitor cystatin C at neutral pH. The resulting modified inhibitor had size and amino acid composition consistent with a cystatin C molecule devoid of the N-terminal Ser-1-Val-10 decapeptide. Leucocyte-elastase-modified cystatin C had more than 240-fold lower affinity than native cystatin C for papain. Removal of the N-terminal decapeptide of human cystatin C also decreased inhibition of human cathepsins B and L by three orders of magnitude, but decreased inhibition of cathepsin H by only 5-fold. A tripeptidyldiazomethane analogue of of the N-terminal portion of cystatin C was a good inhibitor of cathepsins B and L but a poor inhibitor of cathepsin H. It therefore appears that amino acid side chains of the N-terminal segment of cystatin C bind in the substrate-binding pockets of cathepsins B and L but not in those of cathepsin H. It is argued that the N-terminal cystatin C interaction with cathepsin B is physiologically important and hence that leucocyte elastase could have a function as a regulator of extracellular cysteine-proteinase inhibitory activity at sites of inflammation.
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PMID:Human cystatin C. role of the N-terminal segment in the inhibition of human cysteine proteinases and in its inactivation by leucocyte elastase. 199 59

Determination of the primary structure of abnormal Hbs on the basis of DNA sequencing of the globin gene obtained from a carrier of abnormal Hb was performed. DNA obtained from the leukocytes of the peripheral blood was amplified by the polymerase chain reaction (PCR) using the proper amplification primer set. Amplified DNA was digested with two different restriction endonucleases and cloned to vector M 13 mp 18 or mp 19, which had been digested with the same enzymes. DNA sequencing was done by the dideoxy chain termination method using T 7 DNA polymerase, and the abnormal Hbs whose primary structure was determined were as follows: Hb Fukuoka [beta 2 His(CAC/T)----Tyr(TAT)], Hb Machida [beta 6 Glu(GAG)----Gln (CAG)], Hb Hope [beta 136 Gly(GGT)----Asp(GAT)], Hb Hiroshima [beta 146 His(CAC)----Asp(GAC)] and Hb Kodaira [beta 146 His(CAC)----Gln(CAA)]. This method for determining the primary structure of abnormal Hbs might be more effective than the ordinary method, which involves amino acid analysis and amino acid sequencing of the abnormal peptide obtained from abnormal Hb.
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PMID:[Structural analysis of abnormal hemoglobin by the polymerase chain reaction (PCR) of genomic DNA]. 223 67

Hydropathic anticomplementarity of amino acids indicates that peptides derived from complementary DNA strands may form amphiphilic structures and bind one another. By using this concept, we have found that the antisense peptide Ser-Tyr-Asp-Leu complementary to the segment Gln-Ile-Val-Ala-Gly (residues 55-59) in cystatin C (an inhibitor of cysteine proteases) is located at positions 611-614 of the beta chain of human C4, the fourth component of complement. Here we describe and characterize the specific interaction between cystatin C and C4 by ligand affinity chromatography and ELISA. Interaction between the two native proteins was mimicked on replacement of one of them with the corresponding sense-antisense peptide coupled to a carrier protein, and the binding was inhibited by these synthetic peptides in solution. Through the interaction with C4, cystatin C may play a regulatory role in complement activation that might be of particular importance at tissue sites where both proteins are produced by macrophages.
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PMID:Binding of cystatin C to C4: the importance of sense-antisense peptides in their interaction. 230 99

The isolated amyloid substance in hereditary cystatin C amyloid angiopathy (HCCAA) is mainly composed of a cystatin C variant devoid of the 10 amino terminal amino acid residues of extracellular cystatin C from healthy individuals. We have developed a procedure for protein sequencing directly from agarose gel electropherograms and used this in conjunction with isoelectric focusing to investigate the amino terminal sequence of cerebrospinal fluid (CSF) cystatin C in HCCAA patients. The amino-terminal sequence determined for cystatin C from a HCCAA patient CSF sample, Xaa-Ser-Pro-Gly-Lys-Pro-Pro-Xaa-Leu-Val-Gly-Gly-Pro-Met-Xaa-Ala-Xaa-Val, showed that the protein was not amino-terminally truncated. CSF cystatin C from all nine HCCAA patients investigated was found to have an isoelectric point identical to that of native cystatin C, and the truncated form of cystatin C isolated from amyloid deposits was shown to contribute to less than 1% of the total amount of cystatin C in CSF. The total cysteine proteinase inhibitory capacity of CSF from HCCAA patients was lower than that of CSF from other patients. This decreased CSF inhibitory capacity in HCCAA patients was caused by decreased levels of cystatin C, since the levels of the other two cysteine proteinase inhibitors found in CSF, alpha 2-macroglobulin and kininogen, were significantly higher than in CSF from non-HCCAA patients.
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PMID:The amino terminal portion of cerebrospinal fluid cystatin C in hereditary cystatin C amyloid angiopathy is not truncated: direct sequence analysis from agarose gel electropherograms. 231 47

When an excess of human cystatin C or chicken cystatin was mixed with papain, an enzyme-inhibitor complex was formed immediately. The residual free cystatin was then progressively converted to a form with different electrophoretic mobility and chromatographic properties. The modified cystatins were isolated and sequenced, showing that there had been cleavage of a single peptide bond in each molecule: Gly11-Gly12 in cystatin C, and Gly9-Ala10 in chicken cystatin. The residues Gly11 (cystatin C) and Gly9 (chicken cystatin) are among only three residues conserved in all known sequences of inhibitory cystatins. The modified cystatins were at least 1000-fold weaker inhibitors of papain than the native cystatins. An 18-residue synthetic peptide corresponding to residues 4-21 of cystatin C did not inhibit papain but was cleaved at the same Gly-Gly bond as cystatin C. When iodoacetate or L-3-carboxy-trans-2,3-epoxypropionyl-leucylamido-(4-guanidin o)butane was added to the mixtures of either cystatin with papain, modification of the excess cystatin was blocked. Papain-cystatin complexes were stable to prolonged incubation, even in the presence of excess papain. We conclude that the peptidyl bond of the conserved glycine residue in human cystatin C and chicken cystatin probably is part of a substrate-like inhibitory reactive site of these cysteine proteinase inhibitors of the cystatin superfamily and that this may be true also for other inhibitors of this superfamily. We also propose that human cystatin C and chicken cystatin, and probably other cystatins as well, inhibit cysteine proteinases by the simultaneous interactions with such proteinases of the inhibitory reactive sites and other, so far not identified, areas of the cystatins. The cleavage of the inhibitory reactive site glycyl bond in mixtures of papain with excess quantities of cystatins is apparently due to the activity of a small percentage of atypical cysteine proteinase molecules in the papain preparation that form only very loose complexes with cystatins under the conditions employed and degrade the free cystatin molecules.
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PMID:Identification of the probable inhibitory reactive sites of the cysteine proteinase inhibitors human cystatin C and chicken cystatin. 349 37

Foscarnet (phosphonoformic acid) is a pyrophosphate analog that inhibits the replication of human immunodeficiency virus type 1 (HIV-1) in vitro and in patients with AIDS. HIV-1 resistance to foscarnet has not been reported despite long-term foscarnet therapy of AIDS patients with cytomegalovirus disease. We therefore attempted to select foscarnet-resistant HIV-1 in vitro by serial endpoint passage of virus in 400 microM foscarnet. After 13 cycles of passage in MT-2 cells, virus exhibiting > or = 8.5-fold foscarnet resistance was isolated. The reverse transcriptase (RT) from resistant virions exhibited a similar level of foscarnet resistance in enzyme inhibition assays (approximately 10-fold resistance). Foscarnet-resistant virus showed increased susceptibility to 3'-azido-3'-deoxythymidine (90-fold) and to the HIV-1-specific RT inhibitors TIBO R82150 (30-fold) and nevirapine (20-fold). DNA sequence analysis of RT clones from resistant virus revealed the coexistence of two mutations in all clones: Gln-161 to Leu (CAA to CTA) and His-208 to Tyr (CAT to TAT). Sequence analysis of six clinical HIV-1 isolates showing reduced susceptibility to foscarnet revealed the Tyr-208 mutation in two, the Leu-161 mutation in one, and a Trp-88-to-Ser or -Gly mutation in four isolates. Site-specific mutagenesis and production of mutant recombinant viruses demonstrated that the Leu-161, Ser-88, and Tyr-208 mutations reduced HIV-1 susceptibility to foscarnet 10.5-, 4.3-, and 2.4-fold, respectively, in MT-2 cells. In the crystal structure of HIV-1 RT, the Gln-161 residue lies in the alpha E helix beneath the putative deoxynucleoside triphosphate (dNTP) binding site. The Gln-161-to-Leu mutation may affect the structure of the dNTP binding site and its affinity for foscarnet. The location of the Trp-88 residue in the Beta5a strand of HIV-1 RT suggest that the Ser-88 mutation affects template-primer binding, as do several mutations that affect RT susceptibility to nucleoside analogs.
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PMID:Novel mutations in reverse transcriptase of human immunodeficiency virus type 1 reduce susceptibility to foscarnet in laboratory and clinical isolates. 754 60

Cysteine proteases of the papain family generally exhibit broad P1 specificity. A notable exception is papaya proteinase IV (PPIV), which only accepts Gly at this position. In all other cysteine proteases the S1 subsite residues 23 and 65 (papain numbering) are absolutely conserved as Gly, while in PPIV they are replaced by Glu and Arg, respectively. These differences appear to underlie both PPIV specificity and its resistance to inhibition by cystatins. To test this hypothesis, the equivalent residues (Gly27 and Gly73) in the mammalian cysteine protease cathepsin B were changed to Glu and Arg, respectively. Relative to the wild-type enzyme, the Gly27Glu and Gly73Arg mutants showed a drastic reduction in activity with substrates containing a P1 Arg. In contrast, substrates having a Gly residue in P1 were hydrolyzed effectively. The double mutant (Gly27Glu:Gly73Arg) exhibited no detectable activity against any substrate studied. Inhibition of the Gly73Arg mutant by E-64 [1-(L-trans-epoxysuccinyl-L-leucylamino)-4-guanidinobutane] was found to be similar to that of the wild-type enzyme. In contrast, inhibition by cystatin C exhibited a 20,000-fold reduction. These results demonstrate the dramatic influence of side chains at sequence locations 27 and 73 on the S1 subsite specificity of cysteine proteases.
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PMID:Modification of S1 subsite specificity in the cysteine protease cathepsin B. 777 Apr 53

The interaction between cystatin C variants, in which the evolutionarily conserved Gly-11 residue was substituted by Ala, Glu or Trp, and the cysteine proteinases, papain, ficin, actinidin and cathepsin B, was characterized. The substitutions reduced the affinity of binding in a manner consistent with the Gly residue of the wild-type inhibitor, allowing the N-terminal region to adopt a conformation that was optimal for interaction with target proteinases. Replacement of Gly-11 by Ala resulted in only a 5- to 100-fold reduction in binding affinity. Comparison with the affinities of wild-type cystatin C lacking the N-terminal region indicated that even this small structural change affects the conformation of this region sufficiently to largely abolish its interaction with the weakly binding proteinases, actinidin and cathepsin B. However, the substitution allows interactions of appreciable strength between the N-terminal region and the tightly binding enzymes, papain or ficin. Replacement of Gly-11 with the larger Glu and Trp residues substantially decreased the affinity of binding to all enzymes, from 10(3)- to 10(5)-fold. These substitutions further affect the conformation of the N-terminal region, so that interactions of this region with papain and ficin are also essentially eliminated. The decreased affinities of the three cystatin C variants for papain, ficin and actinidin were due exclusively to increased dissociation rate constants. In contrast, the decreased affinity between cathepsin B and the Ala-11 variant, the only one for which rate constants could be determined with this enzyme, was due almost entirely to a decreased association rate constant. This behaviour is analogous to that observed for forms of cystatin C lacking the N-terminal region and supports the conclusion that the mode of interaction of this region with target proteinases varies with the enzyme as a result of structural differences in the active-site region of the latter.
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PMID:Probing the functional role of the N-terminal region of cystatins by equilibrium and kinetic studies of the binding of Gly-11 variants of recombinant human cystatin C to target proteinases. 788 4


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