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排序方式: 共有93条查询结果,搜索用时 9 毫秒
1.
R M Wright M D Yablonsky Z P Shalita A K Goyal D E Eveleigh 《Applied and environmental microbiology》1992,58(11):3455-3465
Genomic DNA fragments encoding beta-glucosidase activities of the thermophilic actinomycete Microbispora bispora were cloned into Escherichia coli. Transformants expressing beta-glucosidase activity were selected by their ability to hydrolyze the fluorogenic substrate 4-methylumbelliferyl-beta-D-glucoside. Two genes encoding beta-glucosidase activity were isolated and distinguished by restriction analysis, Southern hybridization, and the substrate specificities of the encoded enzymes. One gene, bglB, encoded a beta-glucosidase that was expressed intracellularly in E. coli. It exhibited a molecular mass of approximately 52,000 Da by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (PAGE) and 51,280 Da by nondenaturing gradient PAGE, a pI of 4.6, and temperature and pH optima of 60 degrees C and 6.2, respectively. Cloned BglB showed greater activity against cellobiose than against aryl-beta-D-glucosides and was thermostable, retaining about 70% of its activity after 48 h at 60 degrees C. BglB activity is activated two- to threefold in the presence of 2 to 5% (0.1 to 0.3 M) glucose. The DNA sequence of the 2.2-kb insert carrying bglB has been determined. An open reading frame which codes for a protein of 473 amino acids with a predicted molecular mass of 52,227 Da showed significant homology (40 to 47% identity) with beta-glucosidases from glycosal hydrolase family 1. 相似文献
2.
Mechanisms of DNA oxidation 总被引:1,自引:0,他引:1
Aust AE Eveleigh JF 《Proceedings of the Society for Experimental Biology and Medicine. Society for Experimental Biology and Medicine (New York, N.Y.)》1999,222(3):246-252
Oxidative damage of DNA caused by a variety of chemical and physical agents appears to be linked to cancer. However, it is becoming increasingly clear that endogenous generation of oxidants, such as hydroxyl radical and peroxynitrite, lead to oxidation of DNA, and this may cause cancer in individuals where no obvious exposure to chemical or physical agents known to be carcinogenic has occurred. The mechanisms for generation of these two oxidants in living organisms will be discussed and their reactivities with DNA to produce oxidized products (e.g., 8-oxo-dG) will be presented with special emphasis on the individual characteristics of the generation and reactivity of each oxidant. 相似文献
3.
α-Amylase activity during pullulan production and α-amylase gene analyses of <Emphasis Type="Italic">Aureobasidium pullulans</Emphasis> 总被引:1,自引:0,他引:1
4.
5.
André O. S. Lima Diane F. Davis Gavin Swiatek James K. McCarthy Dinesh Yernool Aline A. Pizzirani-Kleiner Douglas E. Eveleigh 《Molecular biotechnology》2009,42(2):205-215
By applying a directed evolution methodology specific enzymatic characteristics can be enhanced, but to select mutants of
interest from a large mutant bank, this approach requires high throughput screening and facile selection. To facilitate such
primary screening of enhanced clones, an expression system was tested that uses a green fluorescent protein (GFP) tag from
Aequorea victoria linked to the enzyme of interest. As GFP’s fluorescence is readily measured, and as there is a 1:1 molar correlation between
the target protein and GFP, the concept proposed was to determine whether GFP could facilitate primary screening of error-prone
PCR (EPP) clones. For this purpose a thermostable β-glucosidase (BglA) from Fervidobacterium sp. was used as a model enzyme. A vector expressing the chimeric protein BglA-GFP-6XHis was constructed and the fusion protein
purified and characterized. When compared to the native proteins, the components of the fusion displayed modified characteristics,
such as enhanced GFP thermostability and a higher BglA optimum temperature. Clones carrying mutant BglA proteins obtained
by EPP, were screened based on the BglA/GFP activity ratio. Purified tagged enzymes from selected clones resulted in modified
substrate specificity. 相似文献
6.
Douglas E Eveleigh Mary Mandels Raymond Andreotti Charles Roche 《Biotechnology for biofuels》2009,2(1):21-8
This article sets forth a simple cellulase assay procedure. Cellulose is variable in nature, insoluble and resistant to enzymatic
attack. As a result there have been a bevy of bewildering cellulase assays published that yielded irrational results. Certain
protocols focused on the rapidity of the assay while ignoring that only the most readily susceptible cellulose regions were
being hydrolyzed. Other assays simplified the system by using modified soluble substrates and yielded results that bore no
relationship to the real world hydrolysis of insoluble cellulose. In this study Mandels, Andreotti and Roche utilized a common
substrate, Whatman filter paper. Hydrolysis of a 50 mg sample of the paper was followed to roughly 4% degradation, which circumvented
the problems of attack of only the most susceptible zones. This common hydrolysis target range also resulted in some balance
with regard to the interaction of the several cellulase components. The method was subsequently widely adopted. 相似文献
7.
1. Oviposition behaviour and host size ? fitness relationships of a gregarious, idiobiont ectoparasitoid, Elachertus cacoeciae (Howard) (Hymenoptera: Eulophidae), were studied by implanting one fourth‐, fifth‐, and sixth‐instar spruce budworm Choristoneura fumiferana (Clemens) (Lepidoptera: Tortricidae) larva per tree in a stand where the density of the wild C. fumiferana population was low. 2. Development time of E. cacoeciae larvae was quickest on fifth‐instar C. fumiferana larvae, which were the preferred hosts for oviposition. 3. Brood sex ratio (proportion of females) was related positively to increasing C. fumiferana instar, indicating that more females were laid on larger hosts. 4. Parasitoid offspring size increased with increasing C. fumiferana instar and decreased with increasing brood size on smaller hosts. Female but not male size was related positively to increasing brood sex ratio (proportion of females). 5. Under laboratory conditions, parasitoid longevity was related positively to parasitoid size and realised lifetime fecundity, and clutch size was related positively to host size. 6. These results suggest that selection of intermediate‐sized C. fumiferana larvae may be adaptive for E. cacoeciae. 相似文献
8.
Double-Antibody Sandwich Enzyme-Linked Immunosorbent Assay for Cellobiohydrolase I 总被引:1,自引:0,他引:1
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A double-antibody sandwich enzyme-linked immunosorbent assay was developed for quantifying cellobiohydrolase I (CBH I) in crude preparations of the cellulase complex from Trichoderma reesei. The other enzymes (endoglucanase and β-glucosidase) in this complex and other ingredients in culture broth did not interfere with this assay. The antibody configuration that resulted in the highest specificity for the assay of CBH I employed a monoclonal antibody to coat wells in polystyrene plates and peroxidase-labeled polyclonal antibody to detect cellobiohydrolase bound to the immobilized monoclonal antibody. Previously, procedures have not been available for the direct assay of CBH I activity in the presence of the other enzymes in the complex, and current indirect procedures are cumbersome and inaccurate. The direct procedure described here is highly specific for CBH I and useful for quantifying this enzyme in the range of 0.1 to 0.8 μg/ml. 相似文献
9.
M. Morris D. E. Eveleigh S. C. Riggs W. N. Tiffney Jr. 《American journal of botany》1974,61(8):867-870
The nodules on roots of Myrica pensylvanica (bayberry) contain a bacterial endophyte. By using the acetylene reduction technique these plant endophyte associations were shown to be capable of fixing nitrogen. As nodulation was plentiful and fixation vigorous, it is proposed that the success of M. pensylvanica as an early successional plant of dunes and impoverished coastal soils is due in part to the nitrogen-fixing capacity of its nodular association. 相似文献
10.
N G Anderson D D Willis D W Holladay J E Caton J W Holleman J W Eveleigh J E Attrill F L Ball N L Anderson 《Analytical biochemistry》1975,66(1):159-174
An automatic system, termed a Cyclum, is described which allows column chromatographic separations to be repeated precisely a large number of times. Provision is made for the adjustment during operation of parameters such as equilibration, wash, elution, and sample flow times and duration of fraction collection. The system is applicable to both analytical and preparative use in various types of column chromatography (e.g., affinity, gel filtration, ion-exchange), but has been especially developed for separations based on immunosorption. 相似文献