| 1. |
- Wahlin, Anders, et al.
(författare)
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Effects of plasmapheresis on the plasma concentration of proteins used to monitor the disease process in multiple myeloma
- 1988
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Ingår i: Acta Medica Scandinavica. - : Wiley. - 0001-6101. ; 223:3, s. 263-267
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Tidskriftsartikel (refereegranskat)abstract
- We studied the influence of plasmapheresis on the plasma concentrations of proteins (IgG, IgA, beta 2-microglobulin) used for estimation of tumour cell mass in patients with multiple myeloma. Simultaneously, the effects of plasmapheresis on plasma concentrations of proteins (CRP, alpha 1-antitrypsin, haptoglobin) used for the assessment of inflammatory processes were studied. Also, changes in the plasma concentration of protein HC, a recently described protein occurring both as a free protein and as an IgA complex, were studied. The influence of plasmapheresis varied for the different proteins. The plasma levels of IgG, IgA, CRP, alpha 1-antitrypsin, and haptoglobin decreased during plasmapheresis. The simultaneous reduction of the level of protein HC was smaller than that of IgG, IgA and haptoglobin. The plasma concentration of beta 2-microglobulin did not decrease during plasmapheresis. The recovery rates of the proteins were found to be approximately inversely related to their molecular weights.
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| 2. |
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| 3. |
- Abrahamson, Magnus, et al.
(författare)
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Efficient production of native, biologically active human cystatin C by Escherichia coli
- 1988
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Ingår i: FEBS Letters. - 1873-3468. ; 236:1, s. 14-18
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Tidskriftsartikel (refereegranskat)abstract
- A cDNA encoding the mature human cysteine proteinase inhibitor cystatin C was fused to the coding sequence for the Escherichia coli outer membrane protein A signal peptide, and the recombinant gene was expressed in E. coli under the control of the λ PR promoter, an optimized Shine-Dalgarno sequence and the λ cI 857 repressor. When induced at 42°C, such cells expressed large amounts of recombinant cystatin C. The recombinant protein was isolated in high yield and characterized. All physicochemical properties investigated, including the positions of disulfide bonds, indicated that the E. coli derived cystatin C was identical to cystatin C isolated from human biological fluids, except that the proline residue in position three was not hydroxylated. The recombinant protein displayed full biological activity against papain, cathepsin B and dipeptidyl peptidase I.
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| 4. |
- Abrahamson, Magnus, et al.
(författare)
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Identification of the probable inhibitory reactive sites of the cysteine proteinase inhibitors human cystatin C and chicken cystatin
- 1987
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Ingår i: Journal of Biological Chemistry. - 1083-351X. ; 262:20, s. 9688-9694
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Tidskriftsartikel (refereegranskat)abstract
- 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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| 5. |
- Abrahamson, Magnus, et al.
(författare)
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Isolation of six cysteine proteinase inhibitors from human urine. Their physicochemical and enzyme kinetic properties and concentrations in biological fluids
- 1986
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Ingår i: Journal of Biological Chemistry. - 1083-351X. ; 261:24, s. 11282-11289
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Tidskriftsartikel (refereegranskat)abstract
- Six cysteine proteinase inhibitors were isolated from human urine by affinity chromatography on insolubilized carboxymethylpapain followed by ion-exchange chromatography and immunosorption. Physicochemical and immunochemical measurements identified one as cystatin A, one as cystatin B, one as cystatin C, one as cystatin S, and one as low molecular weight kininogen. The sixth inhibitor displayed immunochemical cross-reactivity with salivary cystatin S but had a different pI (6.85 versus 4.68) and a different (blocked) N-terminal amino acid. This inhibitor was tentatively designated cystatin SU. The isolated inhibitors accounted for nearly all of the cysteine proteinase inhibitory activity of the urinary pool used as starting material. The enzyme inhibitory properties of the inhibitors were investigated by measuring inhibition and rate constants for their interactions with papain and human cathepsin B. Antisera raised against the inhibitors were used in immunochemical determinations of their concentrations in several biological fluids. The combined enzyme kinetic and concentration data showed that several of the inhibitors have the capacity to play physiologically important roles as cysteine proteinase inhibitors in many biological fluids. Cystatin C had the highest molar concentration of the inhibitors in seminal plasma, cerebrospinal fluid, and milk; cystatin S in saliva and tears; and kininogen in blood plasma, synovial fluid, and amniotic fluid.
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| 6. |
- Abrahamson, Magnus, et al.
(författare)
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Molecular cloning and sequence analysis of cDNA coding for the precursor of the human cysteine proteinase inhibitor cystatin C
- 1987
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Ingår i: FEBS Letters. - : Wiley. - 1873-3468 .- 0014-5793. ; 216:2, s. 229-233
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Tidskriftsartikel (refereegranskat)abstract
- Recombinant cystatin C producing clones were isolated from a human placenta λgt11 cDNA library. The cDNA insert of one of the clones, containing 777 base pairs, encodes the complete mature cystatin C (120 amino acids) and a hydrophobic leader sequence of 26 amino acids, indicating an extracellular function of the inhibitor. The deduced protein sequence confirms the protein sequence of cystatin C isolated from human urine, but differs in one position from the sequence of the cystatin C fragment deposited as amyloid in hereditary cerebral hemorrhage with amyloidosis.
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| 7. |
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| 8. |
- Barrett, Alan, et al.
(författare)
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The place of human gamma-trace (cystatin C) amongst the cysteine proteinase inhibitors
- 1984
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Ingår i: Biochemical and Biophysical Research Communications. - 1090-2104. ; 120, s. 631-636
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Tidskriftsartikel (refereegranskat)abstract
- Native γ-trace, a small basic protein present in high concentration in cerebrospinal fluid, semen and neuroendocrine cells, but of unknown biological function, is shown to be a potent inhibitor of the cysteine proteinases papain, ficin, and human cathepsins B, H and L. It proves to be the tightest-binding protein inhibitor of cathepsin B so far discovered. The name cystatin C is proposed for γ-trace to reflect the many similarities in activity and structure to chicken egg-white cystatin and mammalian cystatins A and B. The inhibition constants of cystatin C, taken together with its widespread distribution in human tissues and extracellular fluids, suggest that a physiological function could well be the regulation of cysteine proteinase activity.
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| 9. |
- Björck, Lars, et al.
(författare)
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Bacterial growth inhibited by a synthetic inhibitor based upon the structure of a human proteinase inhibitor
- 1989
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Ingår i: Nature. - : Springer Science and Business Media LLC. - 0028-0836 .- 1476-4687. ; 337:6205, s. 385-386
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Tidskriftsartikel (refereegranskat)abstract
- Cysteine proteinases are important not only in the intracellular catabolism of peptides and proteins1 and in the processing of prohormones and proenzymes2,3, but also in the penetration of normal human tissue by malignant cells4 and possibly microorganisms5, including viruses. Cystatin C is a human cysteine proteinase inhibitor present in extracellular fluids6. We have synthesized peptide derivatives mimicking the proposed proteinase-binding centre of cystatin C7 and find that they irreversibly inhibit cysteine proteinases. Several bacteria produce proteinases, so we tested a tripeptide derivative (Z-LVG-CHN2) for in vitro anti-bacterial activity against a large number of bacterial strains belonging to thirteen different species. It was found to inhibit specifically the growth of all strains of group A streptococci. The susceptibility of these human pathogens to the peptide was compared with that to well-established anti-streptococcal antibiotics such as tetracy-cline and bacitracin. Moreover, the peptide was active in vivo against group A streptococci: mice injected with lethal doses of these bacteria were cured by a single injection of Z-LVG-CHN2. The cysteine proteinase produced by group A streptococci was isolated and found to be inhibited by Z-LVG-CHN2; moreover, excess proteinase relieved the growth inhibition caused by the peptide derivative, suggesting that the antibacterial activity of Z-LVG-CHN2 is due to inhibition of this cysteine proteinase. This strategy of blocking proteinases with peptide derivatives that mimic naturally occurring inhibitors could be useful in the construction of new agents against other microorganisms, including viruses.
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| 10. |
- Dalboge, H, et al.
(författare)
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High-level expression of active human cystatin C in Escherichia coli
- 1989
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Ingår i: Gene. - 1879-0038. ; 79:2, s. 325-332
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Tidskriftsartikel (refereegranskat)abstract
- Expression of the human cysteine proteinase inhibitor, cystatin C (CysC) in the cytoplasm of Escherichia coli was studied using a cDNA fragment encoding the cysteine proteinase inhibitor controlled by the phage λ pImage /cI857 system. The yield of CysC was low, probably due to proteolytic degradation. By fusing the cysC cDNA to a DNA fragment encoding the signal peptide of the E. coli outer membrane protein A, it was possible to produce a substantial amount of CysC in the periplasm. The processing of the signal peptide was shown to be quantitative and to result in CysC with the correct N-terminal amino acid. Yields higher than 1000 μg CysC/ml can be obtained by initiating the product formation at a moderate temperature (40 °C) late in an optimized fermentation process. A method that gives selective extraction of the periplasmic proteins and at the same time stabilizes CysC has been used.
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