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991.
Helland R Czapinska H Leiros I Olufsen M Otlewski J Smalås AO 《Journal of molecular biology》2003,333(4):845-861
Crystal structures of P1 Gly, Val, Leu and Phe bovine pancreatic trypsin inhibitor (BPTI) variants in complex with two serine proteinases, bovine trypsin and chymotrypsin, have been determined. The association constants for the four mutants with the two enzymes show that the enlargement of the volume of the P1 residue is accompanied by an increase of the binding energy, which is more pronounced for bovine chymotrypsin. Since the conformation of the P1 side-chains in the two S1 pockets is very similar, we suggest that the difference in DeltaG values between the enzymes must arise from the more polar environment of the S1 site of trypsin. This results mainly from the substitutions of Met192 and Ser189 observed in chymotrypsin with Gln192 and Asp189 present in trypsin. The more polar interior of the S1 site of trypsin is reflected by a much higher order of the solvent network in the empty pocket of the enzyme, as is observed in the complexes of the two enzymes with the P1 Gly BPTI variant. The more optimal binding of the large hydrophobic P1 residues by chymotrypsin is also reflected by shrinkage of the S1 pocket upon the accommodation of the cognate residues of this enzyme. Conversely, the S1 pocket of trypsin expands upon binding of such side-chains, possibly to avoid interaction with the polar residues of the walls. Further differentiation between the two enzymes is achieved by small differences in the shape of the S1 sites, resulting in an unequal steric hindrance of some of the side-chains, as observed for the gamma-branched P1 Leu variant of BPTI, which is much more favored by bovine chymotrypsin than trypsin. Analysis of the discrimination of beta-branched residues by trypsin and chymotrypsin is based on the complexes with the P1 Val BPTI variant. Steric repulsion of the P1 Val residue by the walls of the S1 pocket of both enzymes prevents the P1 Val side-chain from adopting the most optimal chi1 value. 相似文献
992.
Modulation of CD40L antigen expression in Jurkat cells: involvement of protein kinase C activity 总被引:1,自引:0,他引:1
Lisowska K Bryl E Soroczyńska M Witkowski JM 《Folia histochemica et cytobiologica / Polish Academy of Sciences, Polish Histochemical and Cytochemical Society》2003,41(4):233-235
The CD40L expressed on activated CD4+ T cells delivers contact-dependent proliferative and anti-apoptotic signals to B lymphocytes. Little is known about molecular mechanisms of constitutive expression of CD40L on some non-Hodgkin's lymphomas, especially about involvement of two signal pathways regulating its expression in normal cells; one involving calcineurin, and the other protein kinase C. We analyzed by flow cytometry the effects of 6-hour stimulation of both pathways (stimuli: PMA and ionomycin) and their inhibitors: cyclosporin A and chelerythrine, on CD40L expression. Two Jurkat clones differing in CD40L surface expression: clone 217.6 (CD40L-) and 217.7 (CD40L+) were studied. Our experiments showed that high level of CD40L expression on the surface of 217.7 cells was reduced after stimulation with PMA. The same effect was observed for combination of PMA and chelerythrine or for PKC inhibitor alone. In 217.6 cells, only chelerythrine used alone induced low level of CD40L expression, while PMA and ionomycin were without effect. These results suggest that CD40L surface expression is mainly dependent on protein kinase C activity. By using PepTag Assay we have confirmed that in both Jurkat clones PKC activity is higher than in normal blood lymphocytes. 相似文献
993.
Lipiński D Jura J Kalak R Pławski A Kala M Szalata M Jarmuz M Korcz A Słomska K Jura J Gronek P Smorag Z Pieńkowski M Słomski R 《Journal of applied genetics》2003,44(2):165-174
The gene construct WAP(6xHisThr):hGH containing the entire human growth hormone gene (hGH) under the rat whey acidic protein (WAP) promoter regulating the expression in mammary glands of mammals was prepared. The 5' end of the gene was modified by the addition of a sequence encoding six histidine residues and a sequence recognized by thrombin. The gene construct was introduced by microinjection into the male pronucleus of a fertilized oocyte. The founder male rabbit was obtained with the transgene mapping to chromosome 7. The presence of the growth hormone was confirmed in samples of milk collected during the lactation of F1 generation females. The growth hormone can be easily purified by affinity chromatography and cleavage by thrombin to an active form. 相似文献
994.
This paper expands the available methods for preparation of H-phosphonoselenoate using a new reagent, 9-fluorenemethyl H-phosphonoselenoate. 相似文献
995.
Bollmark M Johansson T Kullberg M Nilsson J Stawinski J Cieslak J Jankowska J Sobkowski M Szymczak M Wenska M Kraszewski A 《Nucleosides, nucleotides & nucleic acids》2003,22(5-8):617-621
In this paper a short account of our recent research concerning the development of new synthetic methods and reagents for the preparation of nucleotides and their analogues, is given. 相似文献
996.
Exposure of agricultural workers to airborne microorganisms and endotoxin during handling of various vegetable products 总被引:1,自引:0,他引:1
Jacek?DutkiewiczEmail author Ewa?Krysińska-Traczyk Czes?awa?Skórska Jolanta?Sitkowska Zofia?Pra?mo Barbara?Urbanowicz 《Aerobiologia》2000,16(2):193-198
Airborne concentrations of viablemicroorganisms and bacterial endotoxin were determinedduring 10 intramural agricultural activities includinghandling or processing of 6 kinds of vegetableproducts: grain, hay, horticulture seeds, herbs, flaxand potatoes. The median values of the concentrationof total viable microorganisms (bacteria + fungi)were within a range from 43.2 × 103 CFU/m3at potato processing to 1240.0 × 103 CFU/m3in small granaries, exceeding in 9 out of 10 cases thelevel of 105 CFU/m3 suggested as aoccupational exposure limit (OEL). Many of theisolated bacteria and fungi were reported asallergenic and/or immunotoxic agents. The medianvalues of the endotoxin concentration ranged from0.0125 g/m3 at handling hay to54.9 g/m3 at crushing grain, exceeding in 5out of 7 cases the suggested OEL of0.1 g/m3. The high levels of exposure toairborne microorganisms and endotoxin found by thisstudy indicate a potential risk of occupationalrespiratory disorders among agricultural workers, mostly in those handling grain. 相似文献
997.
Witkowski JM Soroczyńska-Cybula M Bryl E Smoleńska Z Jóźwik A 《Journal of immunology (Baltimore, Md. : 1950)》2007,178(2):771-777
Human CD4(+) T lymphocytes undergo aging-related changes leading to decreased immunity to infections and neoplasms, and to increased frequency of autoimmune diseases including rheumatoid arthritis (RA). Certain changes, observed in the CD4(+) cells of RA patients, resemble those observed during physiological aging, but occur at earlier age. Underlying cellular mechanism(s) of these similarities are so far largely unknown. Here we show that KLOTHO, a beta-glucuronidase gene whose activity changes are associated with aging phenotype, is down-regulated at the mRNA, protein, and enzymatic (beta-glucuronidase) activity levels both in the healthy elderly and especially in RA CD4(+) lymphocytes. Although the exact role of Klotho activity for CD4(+) cell function is unknown, we propose here that it might be involved in anti-inflammatory processes occurring in the young and healthy individuals, but reduced in both healthy elderly and RA patients. To support this hypothesis, we show here that the reduction of Klotho expression and activity in both elderly and patients' lymphocytes occurs in concert with the down-regulation of T cell costimulatory molecule CD28, the latter known to be dependent on increased levels of TNF-alpha. Thus, a common mechanism of KLOTHO down-regulation, but executed at various times in life, may underlie both physiological and disease-related T cell aging. Klotho activity might become a target of anti-RA drug development as well as a tool to help increase the immune system efficiency in the elderly. 相似文献
998.
Wu Z Li X Ericksen B de Leeuw E Zou G Zeng P Xie C Li C Lubkowski J Lu WY Lu W 《Journal of molecular biology》2007,368(2):537-549
Human neutrophil alpha-defensins (HNPs) are synthesized in vivo as inactive precursor proteins, i.e. preproHNPs. A series of sequential proteolytic events excise the N-terminal inhibitory pro peptide, leading to defensin maturation and storage in azurophilic granules. The anionic pro peptide, required for correct sub-cellular trafficking and sorting of proHNPs, inhibits the antimicrobial activity of cationic defensins, either inter or intra-molecularly, presumably through charge neutralization. To better understand the role of the pro peptide in the folding and functioning of alpha-defensins and/or pro alpha-defensins, we chemically attached the proHNP1 pro peptide or (wt)pro peptide and the following artificial pro segments to the N terminus of HNP1: polyethylene glycol (PEG), Arg(10) (polyR), Ser(10) (polyS), and (cr)pro peptide, a charge-reversing mutant of the pro peptide where Arg/Lys residues were changed to Asp, and Asp/Glu residues to Lys. Comparative in vitro folding suggested that while all artificial pro segments chaperoned defensin folding, with PEG being the most efficient, the pro peptide catalyzed the folding of proHNPs likely through two independent mechanisms: solubilization of and interaction with the C-terminal defensin domain. Further, the N-terminal artificial pro segments dramatically altered the bactericidal activity of HNP1 against both Escherichia coli and Staphylococcus aureus. Surprisingly, (cr)pro peptide and (wt)pro peptide showed similar properties with respect to intra-molecular and inter-molecular catalysis of defensin folding as well as alpha-defensin binding, although their binding modes appeared different. Our findings identify a dual chaperone activity of the pro peptide and may shed light on the molecular mechanisms by which pro alpha-defensins fold in vivo. 相似文献
999.
Realistic molecular models of one and two-centre catalytic active sites originating from the cleavage of a precursor material
known to give rise to an active double metal cyanide catalyst are described. Via periodic density functional calculations
the structure of the proposed catalytic sites are shown to be dependent on electrostatic and structural relaxation processes
occurring at the surfaces of the precursor material. It is shown how these effects may be adequately captured by small molecular
models of the active sites. The general methodology proposed should provide a computationally efficient basis for detailed
future studies into catalytic reactions over double metal cyanide materials.
Figure Reconstructed DMC [100]-surface: electrostatic potential mapped on charge density isosurface
This work has been originally presented on the Modelling and Design of Molecular Materials conference in Wrocław, Poland. 相似文献
1000.
Aquatic Ecology - Amur sleeper (Perccottus glenii) is currently one of the most invasive fish species in Europe. This inspired our research to identify potential channels of emigration in two... 相似文献