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1.
W. Bockelmann V. Monnet A. Geis M. Teuber J. C. Gripon 《Applied microbiology and biotechnology》1989,31(3):278-282
Summary Cell wall-associated proteinases were isolated from Lactococcus lactis subsp. cremoris AC1 and subsp. lactis NCDO 763 in order to compare their specificities towards different caseins. Two purification strategies were applied. Cells grown in casein-free M17 medium were a suitable starting material for purification, since electrophoretic purity could be achieved after one chromatographic step. Both enzymes has an apparent molecular mass of about 145000 daltons as judged by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Electrophoresis and reversed phase HPLC patterns of hydrolysates of s1-, s2-, -, and K-caseins indicated that both proteinases had a similar specificity. The enzyme of L. lactis subsp. lactis split s1- and s2-caseins more extensively than that of L. lactis subsp. cremoris. 相似文献
2.
Dr. M. E. Stoeckel S. Schimchowitsch J. C. Garaud G. Schmitt H. Vaudry M. J. Klein A. Porte 《Cell and tissue research》1985,242(2):365-370
Summary The immunogold technique, employing antisera with clear-cut specificities, was used to localise different processing stages of pro-opiomelanocortin (POMC) in rabbit melanotropic cells. While the antiserum against
3-MSH labelled all the secretory granules including intrasaccular condensations in the Golgi apparatus, antisera against -MSH only labelled extra-Golgi secretory vesicles (SV). All extra-Golgi SV were likewise labelled with the three antisera against -MSH used, despite their different specificities for the desacetylated, N-acetylated or C-amidated forms of the peptide. The antibody against -endorphin also labelled the extra-Golgi SV, while only some SV were labelled with the antibody against -endorphin. These results correlate with biochemical data in favour of mainly — if not exclusively — intragranular processing of POMC. Except for 3-MSH, the cleavage of which could coincide with Golgi packaging of secretory material, other post-translational modifications of the precursor seem to occur when SV are discharged outside the Golgi area. The cleavage of -endorphin appears to be a later step in POMC processing, occurring in some mature SV. 相似文献
3.
M Watanabe K Morimoto SM Houten N Kaneko-Iwasaki T Sugizaki Y Horai C Mataki H Sato K Murahashi E Arita K Schoonjans T Suzuki H Itoh J Auwerx 《PloS one》2012,7(8):e38286
Background
Besides well-established roles of bile acids (BA) in dietary lipid absorption and cholesterol homeostasis, it has recently become clear that BA is also a biological signaling molecule. We have shown that strategies aimed at activating TGR5 by increasing the BA pool size with BA administration may constitute a significant therapeutic advance to combat the metabolic syndrome and suggest that such strategies are worth testing in a clinical setting. Bile acid binding resin (BABR) is known not only to reduce serum cholesterol levels but also to improve glucose tolerance and insulin resistance in animal models and humans. However, the mechanisms by which BABR affects glucose homeostasis have not been established. We investigated how BABR affects glycemic control in diet-induced obesity models.Methods and Findings
We evaluated the metabolic effect of BABR by administrating colestimide to animal models for the metabolic syndrome. Administration of BABR increased energy expenditure, translating into significant weight reduction and insulin sensitization. The metabolic effects of BABR coincide with activation of cholesterol and BA synthesis in liver and thermogenesis in brown adipose tissue. Interestingly, these effects of BABR occur despite normal food intake and triglyceride absorption. Administration of BABR and BA had similar effects on BA composition and thermogenesis, suggesting that they both are mediated via TGR5 activation.Conclusion
Our data hence suggest that BABR could be useful for the management of the impaired glucose tolerance of the metabolic syndrome, since they not only lower cholesterol levels, but also reduce obesity and improve insulin resistance. 相似文献4.
Dombrink-Kurtzman MA 《Mycopathologia》2003,156(4):357-364
Fumonisins, a family of mycotoxins produced by Fusarium verticillioides (synonym Fusarium moniliforme Sheldon) and F. proliferatum, have been associated with various deleterious effects in different animal species. Serological, hematological and pathological effects and mortality have previously been observed in broiler chicks fed F. proliferatum culture material containing known concentrations of fumonisin, moniliformin and beauvericin. Turkey peripheral blood lymphocytes were exposed in vitro for 72 hours to fumonisin B1(FB1), fumonisin B2(FB2), hydrolyzed fumonisin B1 (HFB1), moniliformin and tricarballylic acid (TCA) (0.01-25 g/ml). A decrease in cell proliferation, as determined by the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] bioassay, occurred in the order: FB2 > FB1 > HFB1, with IC50 = 0.6 M, 1 M and 10 M, respectively. Internucleosomal DNA fragmentation and morphological features characteristic of apoptosis were observed following exposure to fumonisin B1 and beauvericin; cytoplasmic condensation and membrane blebbing were seen by light microscopy. Tricarballylic acid and moniliformin did not interfere with cell proliferation. Results suggested that fumonisin B1 and beauvericin may affect immune functions by suppressing proliferation and inducing apoptosis of lymphocytes. 相似文献
5.
Bearne SL White RL MacDonnell JE Bahrami S Grønlund J 《Molecular and cellular biochemistry》2001,221(1-2):117-126
Methylaspartase (EC 4.3.1.2) was purified 20fold in 35% yield from Fusobacterium varium, an obligate anaerobe. The purification steps included heat treatment, fractional precipitation with ammonium sulfate and ethanol, gel filtration, and ion exchange chromatography on DEAESepharose. The enzyme is dimeric, consisting of two identical 46 kDa subunits, and requires Mg2+ (Km = 0.27 ± 0.01 mM) and K+ (Km = 3.3 ± 0.8 mM) for maximum activity. Methylaspartasecatalyzed addition of ammonia to mesaconate yielded two diastereomeric amino acids, identified by HPLC as (2S,3S)3methylaspartate (major product) and (2S,3R)3methylaspartate (minor product). Optimal activity for the deamination of (2S,3S)3methylaspartate (Km = 0.51 ± 0.04 mM) was observed at pH 9.7. The Nterminal protein sequence (30 residues) of the F. varium enzyme is 83% identical to the corresponding sequence of the clostridial enzyme. 相似文献
6.
通过化学方法使条斑紫菜R-藻红蛋白与抗体进行交联以制备荧光探针, 并对制备条件进行优化。首先采用异双功能试剂SPDP (N-琥珀酰亚氨基-3-2-吡啶基二硫丙酸醇)和DTT(二硫苏糖醇)分别使 R-PE(R-藻红蛋白)衍生化、IgG(单克隆抗体)巯基化, 其次测定了SPDP与R-PE不同摩尔比对R-PE衍生化的影响、DTT与IgG不同摩尔比对IgG巯基化的影响, 结果表明:SPDP与R-PE的最佳摩尔比为40:1, DTT与IgG的最佳摩尔比为500:1。在此基础上, 建立了R-PE与IgG交联的制备技术, 并应用全波长扫描吸收光谱、电泳分析和荧光显微镜观察等监测和分析技术, 证实了藻红蛋白与抗体已成功交联形成了复合物。 相似文献
7.
Human arylamine N-acetyltransferase 1 (NAT1) has been overexpressed in E. coli as a mutant dihydrofolic acid reductase (DHFR) fusion protein with a thrombin sensitive linker. An initial DEAE anion-exchange chromatography resulted in partial purification of the fusion protein. The fusion protein was cleaved with thrombin, and human rNAT1 was purified with a second DEAE column. A total of 8 mg of human rNAT1 from 2 l of cell culture was purified to homogeneity with this methodology. Arylamine substrate specificities were determined for human rNAT1 and hamster rNAT2. With both NATs, the second order rate constants (kcat/Kmb) for p-aminobenzoic acid (PABA) and 2-aminofluorene (2-AF) were several thousand-fold higher than those for procainamide (PA), consistent with the expected substrate specificities of the enzymes. However, p-aminosalicylic acid (PAS), previously reported to be a human NAT1 and hamster NAT2 selective substrate, exhibits 20-fold higher specificity for hamster rNAT2 (k cat/Kmb3410 M-1 s-1 ) than for human rNAT1 (kcat/Kmb 169.4 M-1 s-1 ). p-aminobenzoylglutamic acid (pABglu) was acetylated 10-fold more efficiently by human rNAT1 than by hamster rNAT2. Inhibition studies of human rNAT1 and hamster rNAT2 revealed that folic acid and methotrexate (MTX) are competitive inhibitors of both the unacetylated and acetylated forms of the enzymes, with KI values in 50–300 range. Dihydrofolic acid (DHF) was a much poorer inhibitor of human rNAT1 than of hamster rNAT2. The combined results demonstrate that human rNAT1 and hamster rNAT2 have similar but distinct kinetic properties with certain substrates, and suggest that folic acid, at least in the non-polyglutamate form, may not have an effect on human NAT1 activity in vivo. 相似文献
8.
The polypeptide structure and assembly of Ly-2/3 heterodimers 总被引:1,自引:0,他引:1
Mild reduction of mature, thymic Ly-2/3 heterodimers of M
r 67 000 resulted in dissociation into three individual polypeptide chains, , , and , of respective M
r values 38000, 35000, and 30000. The and chains were both immunoprecipitated by a monoclonal antibody directed to the Ly-2.1 epitope whereas the Ly-3.1 antibody bound only the chain. The possibility that the and chains of each heterodimer established their interchain links within a labile precursor protein in which a and segments were fused was considered but discounted by the finding that in mice heterozygous for both Ly-2 and Ly-3 loci, the Ly-2 product of one chromosome was not exclusively joined to Ly-3 structures coded by the same chromosome. By utilizing ionic detergents which selectively alter the charge of intrinsic membrane proteins, both Ly-2 and Ly-3 polypeptides were shown to have membrane insertion sites. It is suggested that as a consequence of their likely synthesis on membrane-bound polysomes, newly synthesized Ly-2 and Ly-3 structures accumulate within the same subcellular compartment — the membranes of the rough endoplasmic reticulum. Their elevated concentration within this space may facilitate a low affinity binding interaction between Ly-2 and Ly-3 which is later stabilized by interchain disulfide bond formation.Abbreviations used in this paper BSA
bovine serum albumin
- DOC
sodium deoxycholate
- DTT
dithiothreitol
- HA
hemagglutinin
- HTAB
hexadecyltrimethylammonium bromide
- RER
rough endoplasmic reticulum
- SDS-PAGE
sodium dodecyl sulfate polyacrylamide gel electrophoresis
- TX100
Triton X-100 相似文献
9.
《Glycoconjugate journal》1995,12(5):721-728
The expression of neutral glycosphingolipids (GSLs) and gangliosides was investigated in cryosections of normal mouse skeletal muscle and in muscle of mice with neuromuscular diseases using indirect immunofluorescence microscopy. Transversal and longitudinal sections were immunostained with specific polyclonal antibodies against lactosylceramide, lacto-N-neotetraosylceramide, globoside, GM3(Neu5Ac), GM3(Neu5Gc) and GM1(Neu5Ac) as well as monoclonal anti-Forssman GSL antibody. In normal CBA/J mouse muscle (control) the main immunohistochemically detected ganglioside was GM3(Neu5Ac) followed by moderately expressed GM3(Neu5Gc) and GM1. The neutral GSLs lactosylceramide and globoside were stained with almost identical, high fluorescence intensity. Low amounts of lacto-N-neotetraosylceramide and trace quantities of Forssman GSL were immunostained. All GSLs were detected in the sarcolemma, but also in considerable amounts at the intracellular level. Mice with neuromuscular diseases were the A2G-adr mouse mutant (a model for human recessive myotonia of Becker type), the BL6-wr mutant (a model for motor neuron disease) and the BL10-mdx mouse mutant (a model for human Duchenne muscular dystrophy). No changes in GSL expression were found in the A2G-adr mouse, while muscle of the BL6-wr mouse showed increased intensity of immunofluorescence in stainings with anti-lactosylceramide and anti-GM3(Neu5Ac) antibodies. Muscle of BL10-mdx mice showed the most prominent changes in GSL expression with reduced fluorescence intensity for all antibodies. Major differences were not observed in the intensities of GSLs, but there were significant differences in the patterns of distribution on plasma membrane and at the subcellular level. The exact nature and pathogenesis of these changes should be elucidated since such investigations could furnish advances in understanding the functional role of neutral GSLs and gangliosides in normal as well as in diseased muscle.
Abbreviations: BSA, bovine serum albumin; DAPI, 4, 6-diamidine-2-phenylindole-dihydrochloride; DTAF, dichlorotriazinylamino-fluorescein; GSL(s), glycosphingolipid(s); Neu5Ac,N-acetylneuraminic acid; Neu5Gc,N-glycolylneuraminic acid [53]; PBS, phosphate buffered saline. The designation of the following glycosphingolipids follows the IUPAC-IUB recommendations [54] and the nomenclature of Svennerholm [55]. Lactosylceramide or LacCer, Gal1-4Glc1-1Cer; gangliotriaosylceramide or GgOse3Cer, GalNAc1-4Gal1-4Glc1-1Cer; globotriaosylceramide or GbOse3Cer, Gal1-4Gal1-4Glc1-1Cer; gangliotetraosylceramide or GgOse3Cer, Gal1-3GalNAc1-4Gal1-4Glc1-1Cer; globotetraosylceramide or GbOse4Cer, GalNAc1-3Gal1-4Gal1-4Glc1-1Cer; lacto-N-neotetraosylceramide or nLcOse4Cer, Gal1-4GlcNAc1-3Gal1-4Glc1-1Cer; Forssman GSL or GbOse5Cer, GalNAc1-3GalNAc1-3GAl1-4Gal1-4Glc1-1Cer; GM3, II3Neu5Ac-LacCer; GM1, II3Neu5Ac-GgOse4Cer. 相似文献
10.
Ming Zhou Chuner Cai Junxiu Liu Qing Wang Weining Wu Peimin He 《Journal of applied phycology》2010,22(4):459-464
To optimize the chemical conjugation between R-phycoerythrin (R-PE) and antibody, different molar ratios of the heterobifunctional
reagent N-succinimidyl-3-2-pyridyldithio propionate (SPDP) to R-PE were tested for R-PE derivation into R-PE-PDP, and different molar
ratios of dithiothreitol (DTT) to IgG were tested for IgG thiolation. The results showed that in terms of best product yields
determined by ultraviolet (UV) spectrophotometry, the optimal molar ratio of SPDP to R-PE was 40:1 for PE derivation, and
that of DTT to IgG was 500:1 for thiolation. R-PE-labeled secondary antibody was produced by cross-linking PE-PDP and thiolated
IgG. After further purification, UV spectra and native polyacrylamide gel electrophoresis determined its high purity and molecular
weight. Finally, in conjunction with antigen-specific first antibodies, the R-PE-labeled IgG was applied in fluorescence immunoassays
as secondary antibody and successfully detected antigens spotted on nitrocellulose membrane as well as intracellular antigen
in SMCC-7721 cells. This study provides a feasible method of fluorescence antibody preparation from R-PE of Porphyra yezoensis and demonstrates high fluorescent labeling efficiency and good immunologic reactivity of the product. 相似文献
11.
Masaya Ohta Jiharu Hamako Satoru Yamamoto Hajime Hatta Mujo Kim Takehiko Yamamoto Satoru Oka Tsuguo Mizuochi Fumito Matsuura 《Glycoconjugate journal》1991,8(5):400-413
Asparagine-linked oligosaccharides present on hen egg-yolk immunoglobulin, termed IgY, were liberated from the protein by hydrazinolysis. AfterN-acetylation, the oligosaccharides were labelled with a UV-absorbing compound,p-aminobenzoic acid ethyl ester (ABEE). The ABEE-derivatized oligosaccharides were fractionated by anion exchange, normal phase and reversed phase HPLC, and their structures were determined by a combination of sugar composition analysis, methylation analysis, negative ion FAB-MS, 500 MHz1H-NMR and sequential exoglycosidase digestions. IgY contained monoglucosylated oligomannose type oligosaccharides with structures of Glc1-3Man7–9-GlcNAc-GlcNAc, oligomannose type oligosaccharides with the size range of Man5–9GlcNAc-GlcNAc, and biantennary complex type oligosaccharides with core region structure of Man1-6(±GlcNAc1-4)(Man1-3)Man1-4GlcNAc1-4(±Fuc1-6)GlcNAc. The glucosylated oligosaccharides, Glc1Man8GlcNAc2 and Glc1Man7GlcNAc2, have not previously been reported in mature glycoproteins from any source.Abbreviations IgG, IgM, IgD, IgE, and IgA
immunoglobulin G, M, D, E, and A, respectively
- IgY
egg-yolk antibody
- ABEE
p-aminobenzoic acid ethyl ester
- HPLC
high performance liquid chromatography
- FAB-MS
fast atom bombardment mass spectrometry
- Hex
hexose
- HexNAc
N-acetylhexosamine
- hCG
human chorionic gonadotropsin 相似文献
12.
Detailed karyological surveys of the ant Myrmecia pilosula species group, which is characterized by the lowest chromosome number in higher organisms (2n=2), were attempted. We revealed that this species has developed highly complicated chromosomal polymorphisms. Their chromosome numbers are in the range 2n=2, 3, and 4, and six polymorphic chromosomes are involved, i.e., two for chromosome 1 (denoted as SM1 and ST1), three for chromosome 2 (A2, A2, and M2), and M(1+2) for the 2n=2 karyotype. We suggested that these chromosomes were induced from a pseudo-acrocentric (A
1
M
) and A2 as follows: (1) A
1
M
SM1 or ST1 by two independent pericentric inversions; (2) A2A2M2 by chromosomal gap insertion and centromere shift; and (3) ST1+A2M(1+2) by telomere fusion, where (3) means that the 2n=2 karyotype was derived secondarily from a 2n=4 karyotype. It is a noteworthy finding that active nucleolus organizer (NOR) sites, in terms of silver staining, are tightly linked with the centromere in this species, and that both the centromere and NOR of A2 were inactivated after the telomere fusion. 相似文献
13.
Johannes Müthing Jasna Peter-Katalinić Franz-Georg Hanisch Ulrich Neumann 《Glycoconjugate journal》1991,8(5):414-423
YAC-1 cells were propagated in bioreactors in 11 and 7.51 volumes. The cells were metabolically labelled withd-[1-14C]galactose andd-[1-14C]glucosamine. The ganglioside fraction, purified by DEAE-Sepharose and silica gel column chromatography, showed on thin layer chromatography four major bands with mobilities between GM1 and GD1a. Gangliosides, obtained by further purification steps including high performance liquid chromatography on silica gel 60 columns with a gradient system of isopropanol:hexane:water, and preparative high performance TLC were characterized by (1) immunostaining of corresponding asialogangliosides obtained by mild acid hydrolysis and neuraminidase treatment and (2) fast atom bombardment mass spectrometry of native and permethylated samples and methylation analysis of GM1b ganglioside. As well as small amounts of GM2 and GM1, the major gangliosides found in the complex mixture were GM1b and GalNAc-GM1b. The structural heterogeneity of these gangliosides was cased by (a) substitution of the ceramide moiety by fatty acids of different chain length and degree of unsaturation (C16:0, C24:0, C24:1) and (b) N-substitution of the sialic acid moieties with either acetyl or glycolyl groups. Disialogangliosides were detected only in low amounts and will be the subject of further investigation. A polyclonal chicken antiserum was raised against IVNeuAc-GgOse5Cer. The antiserum was highly specific for gangliosides (IVNeuAc and IVNeuGc) and asialogangliosides with a GgOse5Cer backbone. No cross-reaction with GM1b or GgOse4Cer was observed.
Abbreviations: FAB-MS, fast atom bombardment mass spectrometry; GSL(s), glycosphingolipid(s); HPLC, high performance liquid chromatography, HPTLC, high performance thin layer chromatography; NK, natural killer; SIM, selective ion monitoring; TIC, total ion current. NeuAc,N-acetylneuraminic acid; NeuGc,N-glycolylneuraminic acid. The designation of the following glycosphingolipids follows the IUB-IUPAC recommendations. GgOse3Cer or gangliotriaosylceramide or asialo GM2, GalNAc1-4Gal1-4GlcCer; GgOse4Cer or gangliotetraosylceramide or asialo GM1, Gal1-3GalNAc1-4Gal1-4GlcCer; GgOse5Cer organgliopentaosylceramide, GalNAc1-4Gal1-3GalNAc1-4Gal1-4GlcCer; II3NeuAc-GgOse4Cer or GM1; IV3NeuAcGgOse4Cer or GM1b; IV3NeuAc-GgOse5Cer or GalNAc-GM1b; IV3NeuAc, II3NeuAc-GgOse4Cer or GD1a; II3(NeuAc)2-GgOse4Cer or GD1b; IV3(NeuAc)2-GgOse4Cer or GD1c; IV3NeuAc,III6NeuAc-GgOse4Cer or GD1a; IV3NeuAc,II3(NeuAc)2-GgOse4Cer or GT1b;Vibrio cholerae and Arthrobacter ureafaciens neuraminidase (EC 3.2.1.18). 相似文献
14.
Philip H. Johnson Alastair S. R. Donald James Feeney Winifred M. Watkins 《Glycoconjugate journal》1992,9(5):251-264
The acceptor specificity and general properties of a Lewis blood-group gene associated -3/4-L-fucosyltransferase isolated from human milk have been examined at the penultimate purification stage involving affinity chromatography on GDP-hexanolamine Sepharose, and after a subsequent gel filtration step on Sephacryl S-200. Both preparations transferred fucose to theO-4 position ofN-acetylglucosamine in Type 1 (Gal1-3GlcNAc-R) acceptors and theO-3 position of glucose in lactose-based (Gal1-4Glc) oligosaccharides, and both used Type 1 sialylated compounds when the terminalN-acetylneuraminic acid was present in -2,3 linkage. The striking difference between the two preparations was in their reactivity with Type 2 (Gal1-4GlcNAc-R) chains; after Sephacryl S-200 chromatography the apparentK
M values for the -3/4- preparation with unsubstituted low-molecular-weight Type 2 oligosaccharides were considerably increased. Substitution of the terminal galactose with sialic acid in -2,3 linkage decreased theK
M values for low-molecular-weight oligosaccharides but no detectable incorporation of fucose was observed intoN-acetyllactosamine end-groups of glycoproteins withN-linked oligosaccharide chains, irrespective of the presence of sialic acid in the terminal sequences.Deceased 25 June 1991. 相似文献
15.
The four allotypic specificities, z1, z2, y3, and y4, of rabbit
2
-macroglobulin (
2
M) were found to be inherited as three phenogroups, z1y3, z1y4, or z2y3. The phenogroup inheritance can be viewed as reflecting the close linkage of two loci, i.e., the Mtz locus with the z
1 and z
2 alleles and the Mty locus with the y
3 and y
4 alleles. Radioimmunoassay of
2
M by an indirect precipitation method confirmed our earlier studies by a direct method showing that nearly all molecules of z1 or z2 homozygotes possess either z1 or z2, respectively, and that most molecules in the heterozygote are hybrids with both z2 and z2. Unlabeled
2
M from homozygotes and heterozygotes was used to competitively inhibit the reaction of anti-z1 or anti-z2 with labeled
2
M. This revealed that z1z2 heterozygotes possess approximately 35% of the z1 and 64% of the z2 determinants found in the respective homozygotes. These results resemble previous observations with the y3 and y4 allotypes which indicated an unequal contribution of allelic genes in the formation of rabbit
2
M and the lack of allelic exclusion in most of the molecules. Unlike immunoglobulins and low-density lipoproteins, hybrid
2
M molecules form by the interaction of homologous chromosomes.This investigation was supported in part by U.S. Public Health Service Grants AI-09241 and AI-07043. Presented in part at the 55th Annual Meeting of the Federation of American Societies for Experimental Biology, Chicago, Illinois, April 12–17, 1971.In partial fulfillment of the requirements for the Doctor of Philosophy Degree in the Graduate College of the University of Illinois. Supported by a postdoctoral fellowship from the Schweppe Foundation.Recipient of U.S. Public Health Service Research Career Development Award AI-28687. 相似文献
16.
Johannes Müthing Frank Unland Dagmar Heitmann Michaela Orlich Franz-Georg Hanisch Jasna Peter-Katalinić Vera Knäuper Harald Tschesche Sørge Kelm Roland Schauer Jürgen Lehmann 《Glycoconjugate journal》1993,10(1):120-126
The structures of gangliosides from human granulocytes were elucidated by fast atom bombardment mass spectrometry and by gas chromatography/mass spectrometry as their partially methylated alditol acetates. In human granulocytes besides GM3 (II3Neu5Ac-LacCer), neolacto-series gangliosides (IV3Neu5Ac-nLcOse4Cer, IV6Neu5Ac-nLcOse4Cer and VI3Neu5Ac-nLcOse6Cer) containing C24:1, and to some extent C22:0; and C16:0 fatty acid in their respective ceramide portions, were identified as major components. In this study we demonstrate that gangliosides from human granulocytes, the second most abundant cells in peripheral blood, can serve as receptors for influenza viruses A/PR/8/34 (H1N1), A/X-31 (H3N2), and a parainfluenza virus Sendai virus (HNF1, Z-strain). Viruses were found to exhibit specific adhesion to terminal Neu5Ac2-3Gal and/or Neu5Ac2-6Gal sequences as well as depending on the chain length of ganglioside carbohydrate backbones from human granulocytes, these important effector cells which represent the first line of defence in immunologically mediated reactions.
Abbreviations: FAB-MS, fast atom bombardment mass spectrometry; GC/EIMS, gas chromatography/electron impact mass spectrometry; GSL(s) glycosphingolipids; HPTLC, high performance thin-layer chromatography; Neu5Ac,N-acetylneuraminic acid [26], PFU, plaque forming unit. The designation of the following glycosphingolipids follows the IUPAC-IUB recommendations, and the ganglioside nomenclature system of Svennerholm was used. LacCer or lactosylceramide, Gal1-4Glc1-1Cer gangliotetraosylceramide or GgOse4Cer, Gal1-3GalNAc1-4Gal1-4Glc1-1Cer; lacto-N-tetraosylceramide or nLcOse4Cer, Gal1-4GlcNAc1-3Gal1-4-Glc1-1Cer; lacto-N-norhexaosylceramide or nLcOse6Cer, Gal1-4GlcNAc1-3Gal1-4GlcNAc1-3Gal 1-4-Glc1-1Cer; GM3, II3Neu5Ac-LacCer; GM1, II3Neu5Ac-GgOse4Cer; GD1a, IV3Neu5Ac, II3Neu5Ac-GgOse4Cer; GD1b, II3(Neu5Ac)2-GgOse4Cer; GT1b, IV3Neu5Ac, II3(Neu5Ac)2-GgOse4Cer; GQ1b, IV3(Neu5Ac)2, II3(Neu5Ac)2-GgOse4Cer; sialyllacto-N-tetraosylceramide, IV3Neu5Ac/IV6Neu5Ac-nLcOse4Cer; sialyllacto-N-norhexaosylceramide or i-active ganglioside, VI3Neu5Ac-nLcOse6Cer. 相似文献
17.
Coenzyme A-linked acetaldehyde dehydrogenase (ACDH) of ethanol-grown cells of Acetobacterium woodii was purified to apparent homogeneity; a 28-fold purification was achieved with 13% yield. The enzyme proved to be oxygen-sensitive and was inactive in the absence of dithioerythritol. During the purification procedure addition of 1 mM MgCl2 was necessary to maintain enzyme activity. Alcohol dehydrogenase (ADH) activity was separated from ACDH during anion exchange chromatography using DEAE Sephacel. A part of the ACDH activity coeluted with ADH, but both could be separately eluted from a Cibacron Blue 3GA-Agarose column, revealing the same subunit structure and activity band for ACDH as found before and, thus, indicating an aggregation of the enzyme. The remaining ADH activity could be separated by gel filtration. For the native ACDH a molecular mass of 255 kDa was determined by polyacrylamide gel electrophoresis and of 272 kDa by gel filtration using Superose 12. The enzyme subunit sizes were 28 kDa and 40 kDa, respectively, indicating a 44 structure for the active form. The enzyme catalyzed the oxidation of several straight chain aldehydes although it was most active with acetaldehyde. NADH strongly inhibited oxidation of acetaldehyde whereas NADPH had no effect. The inhibition was noncompetitive.Non-standard abbrevations ACDH
acetaldehyde dehydrogenase
- ADH
alcohol dehydrogenase
- CHES
2-(N-cyclohexylamino)-ethanesulfonate
- DTE
dithioerythritol
- KP-buffer
25 mM K-PO4, pH 7.5, containing, 4 mM DTE
- MES
2-(N-morpholino)-ethanesulfonate
- TAPS
N-Tris-(hydroxymethyl)-methyl-3-aminopropa-nesulfonate 相似文献
18.
Interrelationships Between Nitrogen Supply and Photosynthetic Parameters in Vicia faba L. 总被引:2,自引:2,他引:0
We determined for Vicia faba L the influence of nitrogen uptake and accumulation on the values of photon saturated net photosynthetic rate (P
Nmax), quantum yield efficiency (), intercellular CO2 concentration (C
i), and carboxylation efficiency (C
e). As leaf nitrogen content (NL) increased, the converged onto a maximum asymptotic value of 0.0664±0.0049 mol(CO2) mol(quantum)–1. Also, as NL increased the C
i value fell to an asymptotic minimum of 115.80±1.59 mol mol–1, and C
e converged onto a maximum asymptotic value of 1.645±0.054 mol(CO2) m–2 s–1 Pa–1 and declined to zero at a NL-intercept equal to 0.596±0.096 g(N) m–2. fell to zero for an NL-intercept of 0.660±0.052 g(N) m–2. As NL increased, the value of P
Nmax converged onto a maximum asymptotic value of 33.400±2.563 mol(CO2) m–2 s–1. P
N fell to zero for an NL-intercept of 0.710±0.035 g(N) m–2. Under variable daily meteorological conditions the values for NL, specific leaf area (L), root mass fraction (Rf), P
Nmax, and remained constant for a given N supply. A monotonic decline in the steady-state value of Rf occurred with increasing N supply. L increased with increasing N supply or with increasing NL. 相似文献
19.
P Hermentin R Doenges U Franssen C Bieva F J Vander Brugghen P Stryckmans H J Friesen B Optaczy S Schneider W Ax 《Bioconjugate chemistry》1990,1(6):411-418
Iron oxide particles of average size 0.5-1.5 microns, covered by a silane coat carrying amino groups (Bio-Mag, Advanced Magnetics, Boston), were derivatized by reaction with N-[(gamma-maleimidobutyryl)oxy]-succinimide (GMBS), N-hydroxysuccinimidyl iodoacetate (NHIA), 2-iminothiolane (2-It), or N-succinimidyl 3-(2-pyridyldithio)propionate (SPDP). The derivatized particles were suitable for the reaction with sulfhydryl groups and subsequently coated with monoclonal antibodies (MoAbs) of different classes and isotypes (IgM, IgG1, IgG2a, IgG2b, IgG3) as well as polyclonal rabbit anti-mouse IgG (RAM). The antibodies were reduced by dithiothreitol (DTT) and covalently conjugated to the BioMag derivatives via liberated sulfhydryls of the hinge region. The observed conjugation ratios, expressed as protein/iron (micrograms/mg), could be reproducibly varied for optimization. These ratios were dependent on the type and amount of antibody offered for coupling to the derivatized particles, decreasing as follows: polyclonal = IgM greater than IgG2b greater than IgG2a = IgG3 greater IgG1. The conjugation ratios were also dependent on the type and amount of the spacer used to derivatize the BioMag particles, decreasing as follows: GMBS greater than NHIA greater than 2-It greater than SPDP. The magnetically responsive magnetite-antibody conjugates ("magneto-beads"), carrying MoAb BMA 081 (anti-CD8; IgG2a), MoAb BB10 (anti-CD10/CALLA; IgG2b), MoAb VIL-A1 (anti-CD10; IgM), and polyclonal RAM, coupled similarly via 3.6 mumol of GMBS spacer per mg of Fe, were further investigated with respect to a depletion effect on specific cell subsets. The rates of cell depletion were found to be strongly dependent on the individual characteristics of the antibody used.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
20.
M. Wiersma G. Versteegh H. A. Assink G. W. Welling W. Harder 《Antonie van Leeuwenhoek》1978,44(2):157-169
The purification and characterisation of an extracellular endo and aminopeptidase of the marineVibrio SA1 is described. The endopeptidase was purified by ammonium sulphate precipitation, gel filtration and affinity chromatography. It had a molecular weight of approximately 31 000, a pH optimum at 7.8 and a temperature optimum at 50 C. The enzyme was rapidly inactivated at 65 C.The aminopeptidase was purified by ammonium sulphate precipitation, gel filtration and preparative polyacrylamide gel electrophoresis. This enzyme had a molecular weight of approximately 21 000, a pH optimum at 8.6 and a temperature optimum at 60 C. Both proteases were inactivated by EDTA while reactivation occurred by Ca2+, Zn2+ and Mg2+ ions.The endopeptidase hydrolysed several peptide bonds in the oxidized B-chain of insulin, particularly those involving amino groups of hydrophobic amino acid residues with bulky side chains. It was unable to hydrolyse synthetic dipeptides, but a number of tripeptides were hydrolysed at a low rate. The aminopeptidase hydrolysed leucinamide and di- and tripeptides containing hydrophobic bulky amino acids as the N-terminal residue. It was concluded that the endopeptidase and the aminopeptidase ofVibrio SA1 possess complementary specificities.Abbreviations T=
Triethylenetetramine
- S=
Succinic acid
- PIPES=
Piperazine-N,Nbis(2-ethane sulfonic acid)
- HEPES=
N-2-hydroxy-ethylpiperazine-N-2-ethane sulfonic acid
Part of this study was supported by the Foundation for Fundamental Biological Research (BION), which is subsidized by the Netherlands Organization for the Advancement of Pure Research (ZWO). 相似文献