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101.
102.
The catalytic domains of murine Golgi alpha1,2-mannosidases IA and IB that
are involved in N-glycan processing were expressed as secreted proteins in
P.pastoris . Recombinant mannosidases IA and IB both required divalent
cations for activity, were inhibited by deoxymannojirimycin and
kifunensine, and exhibited similar catalytic constants using
Manalpha1,2Manalpha-O-CH3as substrate. Mannosidase IA was purified as a 50
kDa catalytically active soluble fragment and shown to be an inverting
glycosidase. Recombinant mannosidases IA and IB were used to cleave
Man9GlcNAc and the isomers produced were identified by high performance
liquid chromatography and proton-nuclear magnetic resonance spectroscopy.
Man9GlcNAc was rapidly cleaved by both enzymes to Man6GlcNAc, followed by a
much slower conversion to Man5GlcNAc. The same isomers of Man7GlcNAc and
Man6GlcNAc were produced by both enzymes but different isomers of
Man8GlcNAc were formed. When Man8GlcNAc (Man8B isomer) was used as
substrate, rapid conversion to Man5GlcNAc was observed, and the same
oligosaccharide isomer intermediates were formed by both enzymes. These
results combined with proton-nuclear magnetic resonance spectroscopy data
demonstrate that it is the terminal alpha1, 2-mannose residue missing in
the Man8B isomer that is cleaved from Man9GlcNAc at a much slower rate.
When rat liver endoplasmic reticulum membrane extracts were incubated with
Man9GlcNAc2, Man8GlcNAc2was the major product and Man8B was the major
isomer. In contrast, rat liver Golgi membranes rapidly cleaved
Man9GlcNAc2to Man6GlcNAc2and more slowly to Man5GlcNAc2. In this case all
three isomers of Man8GlcNAc2were formed as intermediates, but a distinctive
isomer, Man8A, was predominant. Antiserum to recombinant mannosidase IA
immunoprecipitated an enzyme from Golgi extracts with the same specificity
as recombinant mannosidase IA. These immunodepleted membranes were enriched
in a Man9GlcNAc2to Man8GlcNAc2- cleaving activity forming predominantly the
Man8B isomer. These results suggest that mannosidases IA and IB in Golgi
membranes prefer the Man8B isomer generated by a complementary mannosidase
that removes a single mannose from Man9GlcNAc2.
相似文献
103.
CO(2)-Enhanced Yield and Foliar Deformation among Tomato Genotypes in Elevated CO(2) Environments 总被引:1,自引:0,他引:1
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Yield increases observed among eight genotypes of tomato (Lycopersicon esculentum Mill.) grown at ambient CO2 (about 350) or 1000 microliters per liter CO2 were not due to carbon exchange rate increases. Yield varied among genotypes while carbon exchange rate did not. Yield increases were due to a change in partitioning from root to fruit. Tomatoes grown with CO2 enrichment exhibited nonepinastic foliar deformation similar to nutrient deficiency symptoms. Foliar deformation varied among genotypes, increased throughout the season, and became most severe at elevated CO2. Foliar deformation was positively related to fruit yield. Foliage from the lower canopy was sampled throughout the growing season and analysed for starch, K, P, Ca, Mg, Fe, and Mn concentrations. Foliar K and Mn concentrations were the only elements correlated with deformation severity. Foliar K decreased while deformation increased. In another study, foliage of half the plants of one genotype received foliar applications of 7 millimolar KH2PO4. Untreated foliage showed significantly greater deformation than treated foliage. Reduced foliar K concentration may cause CO2-enhanced foliar deformation. Reduced K may occur following decreased nutrient uptake resulting from reduced root mass due to the change in partitioning from root to fruit. 相似文献
104.
Babesia bovis: gene isolation and characterization using a mung bean nuclease-derived expression library 总被引:9,自引:0,他引:9
Genomic DNA prepared from erythrocyte cultures of Babesia bovis merozoites was digested with mung bean nuclease and used to construct a lambda gt11 expression library of B. bovis recombinants. Immunoscreening with two polyclonal antibody probes detected multiple recombinants from which two, designated Bb-1 and Bb-3, were chosen for further analysis. Monospecific immunoglobulins isolated from the screening sera using nitrocellulose-bound fusion proteins were employed to determine the native molecular weight and the intracellular location of the babesial proteins encoded by the recombinants. Clone Bb-1 encodes an antigen of 77,000 Da located at the apical end of the intraerythrocytic parasite. A protein of 75,000 Da encoded by clone Bb-3 is associated with the infected red blood cell cytoplasm and/or membrane but not with the merozoite. 相似文献
105.
Relationship between sporulation medium and germination ability of Mucor racemosus sporangiospores 总被引:2,自引:0,他引:2
Asexual sporangiospores of Mucor racemosus produced on a minimal sporulation medium (M spores) germinated only if glucose, mannose or a complex substrate such as peptone, yeast extract or Casamino acids was present. Once germinated, growth was supported by a wide range of substrates including amino acids, carbohydrates or organic acids. Sporangiospores produced on a nutritionally complex sporulation medium (C-spores) germinated on a wide range of carbon sources. C-spore phenotype was pleiotropic in that sporangiospores capable of germinating on cellobiose could always germinate on glutamate or xylose; but C-spores capable of germinating on xylose or glutamate did not always germinate on cellobiose. There was a hierarchy of substrates capable of initiating germination with glucose = mannose greater than xylose greater than glutamate greater than cellobiose. C-spores also differed from M-spores by initiating germination in the presence of the non-metabolizable glucose analogue 3-O-methylglucose. These results suggest that at least two sporangiospore phenotypes are produced depending upon the concentration and type of ingredients present in the sporulation medium. 相似文献
106.
107.
108.
Flood JA Tripp TJ Davis CC Hill DR Schlievert PM 《Journal of microbiological methods》2004,57(2):283-288
Human behaviours and consumer products may affect vaginal microbial ecology, thereby influencing women's health. Relevant experimentation systems are needed to understand such possible links. Here, we describe the development of a practical semi-solid in vitro model to assess the effects of interactions between vaginal environment and the presence of tampons, on bacterial communities, including the production of toxic shock syndrome toxin-1 (TSST-1) by Staphylococcus aureus. 相似文献
109.
Bonafoux D Bonar S Christine L Clare M Donnelly A Guzova J Kishore N Lennon P Libby A Mathialagan S McGhee W Rouw S Sommers C Tollefson M Tripp C Weier R Wolfson S Min Y 《Bioorganic & medicinal chemistry letters》2005,15(11):2870-2875
A series of 21 novel 2-[(aminocarbonyl)amino]-5-acetylenyl-3-thiophenecarboxamides were synthesized and evaluated for the inhibition of IKK-2. In spite of their often modest activity on the enzyme, six selected analogs showed significant inhibition of the production of inflammatory cytokine IL-8 in IL-1beta stimulated rheumatoid arthritis-derived synovial fibroblasts, demonstrating their potential usefulness as NF-kappaB regulators. 相似文献
110.
P. J. Fleming P. S. Blair C. Bacon D. Bensley I. Smith E. Taylor J. Berry J. Golding J. Tripp 《BMJ (Clinical research ed.)》1996,313(7051):191-195
OBJECTIVE--To investigate the role of sleeping arrangements as risk factors for the sudden infant death syndrome after a national risk reduction campaign. DESIGN--Two year population based case-control study. Parental interviews were conducted for each infant who died and for four controls matched for age and date of interview. SETTING--Three regions in England with a total population of 17 million people. SUBJECTS--195 babies who died and 780 matched controls. RESULTS--Prone and side sleeping positions both carried increased risks of death compared with supine when adjusted for maternal age, parity, gestation, birth weight, exposure to smoke, and other relevant factors in the sleeping environment (multivariate odds ratio = 9.00 (95% confidence interval 2.84 to 28.47) and 1.84 (1.02 to 3.31), respectively). The higher incidence of side rather than prone sleeping led to a higher population attributable risk (side 18.4%, prone 14.2%). More of the infants who died were found with bed covers over their heads (21.58; 6.21 to 74.99). The use of a dummy had an apparent protective effect (0.38; 0.21 to 0.70). Bed sharing for the whole night was a significant risk factor for infants whose mothers smoked (9.25; 2.31 to 34.02). No protective effect of breast feeding could be identified on multivariate analysis. CONCLUSIONS--This study confirms the importance of certain risk factors for the sudden infant death syndrome and identifies others-for example, covers over the head, side sleeping position-which may be amenable to change by educating and informing parents and health care professionals. 相似文献