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1.
Gabaculine (2,3-dihydro 3-amino benzoic acid) is a potent inhibitor of tetrapyrrole biosynthesis in organisms that use the C5 pathway for the synthesis of δ-aminolaevulinic acid. Glutamate semialdehyde aminotransferase (GSA-AT), the enzyme catalysing the formation of this key precursor of tetrapyrroles, is normally inhibited by concentrations of gabaculine in the order of 5?μM. However, in Synechococcus 6301 strain GR6, a cyanobacterium that is resistant to 100?μM gabaculine, this enzyme has undergone two changes in structure: a deletion of three amino acids from positions 5 to 7 and the substitution of isoleucine for methionine at position 248. To establish the effect in vivo of these specific changes in the gene for GSA-AT (hemL), a suicide vector (pHS7) containing an antibiotic cassette was constructed to achieve the replacement, by homologous recombination, of the wild-type hemL gene in the chromosome by a modified form of the gene. Recombinant strains of Synechococcus 7942 obtained using pHS7-hemL GR6 were indistinguishable from Synechococcus 6301 GR6 in terms of the resistance of growth and of chlorophyll accumulation to high concentrations of gabaculine, while a wild-type recombinant produced using pHS7-hemL WT had retained its sensitivity. Southern hybridisation using gene probes for hemL, amp r and cm r confirmed that chromosomal integration of the plasmids had occurred in both WT and GR6 recombinants. Growth and chlorophyll accumulation in equivalent strains with the hemL gene containing either the deletion or the transition characteristic of Synechococcus 6301 GR6 were inhibited by 10?μM gabaculine. Consequently, resistance in vivo to high concentrations of this compound is dependent on both the changes in gene/enzyme structure. This investigation has established the effectiveness of the suicide vector pHS7 for studying the effect in vivo of specific changes in the hemL gene. It has also demonstrated that replacement of the wild-type gene by that from Synechococcus 6301 GR6 is sufficient to confer resistance in vivo to high concentrations of gabaculine. 相似文献
2.
Cyanobacterial GR6 glutamate-1-semialdehyde aminotransferase: a novel enzyme-based selectable marker for plant transformation 总被引:2,自引:0,他引:2
Angelica Giancaspro Daniele Rosellini Antonio Blanco Agata Gadaleta 《Plant cell reports》2001,20(4):296-300
The mutant glutamate-1-semialdehyde aminotransferase (GSA-AT) enzyme encoded by the hemL gene of the gabaculine-resistant cyanobacterium Synechococcus PCC6301 strain GR6 was expressed in tobacco following Agrobacterium-mediated transformation of leaf discs. When targeted to plastids, the GR6 hemL gene product conveyed gabaculine resistance to transgenic plants. Selection using 50 and 100 µM gabaculine was shown to produce two distinct explant phenotypes: 'greens' and 'whites'. T1 progeny displayed Mendelian segregation ratios, and PCR analysis demonstrated the 'green' phenotype corresponded with the presence of the GR6 hemL gene. Furthermore, 'whites' could be rescued after 9 days growth on solid media containing between 5 µM and 25 µM gabaculine, allowing the potential use of this system for the isolation of gabaculine-sensitive transformants in mutagenesis screening. The use of GR6 hemL as a selectable marker gene provides a novel enzyme-based method for the selection of transgenic regenerants without the need for antibiotic-resistance markers. 相似文献
3.
K. Sangthongpitag R. J. Penfold S. F. Delaney P. L. Rogers 《Applied microbiology and biotechnology》1997,47(4):379-384
Genes encoding the mosquitocidal binary toxin of Bacillus sphaericus 2362 were introduced into Synechococcus PCC6301, a cyanobacterium that can tolerate a number of potential variations in the mosquito breeding environment, and can
serve as a food source for mosquito larvae. The toxin genes, preceded by a Synechococcus rbcL promoter, were located on a mobilizable Escherichia coli Synechococcus shuttle vector, which was introduced into Synechococcus PCC6301 at frequencies of 10−5–10−7 exconjugants/recipient, depending on the selective conditions used. Recombinant Synechococcus exhibited significant toxicity against 2-day-old and 6-day-old Culex quinquefasciatus larvae, the concentration required to kill 50 % of larvae (LC50) being 2.1 × 105 and 1.3 × 105 cells/ml respectively. Mosquitocidal activity decreased tenfold after 20 generations of non-selective growth.
Received: 23 July 1996 / Received revision: 11 November 1996 / Accepted: 15 November 1996 相似文献
4.
Glutamate-l-semialdehyde (GSA) aminotransferase catalyses the final step in the C5 pathway converting glutamate to the tetrapyrrole precursor δ-aminolaevulinic acid. This enzyme is sensitive to gabaculine (2,3-dihydro-3-amino benzoic acid) and to 4-amino-5-fluoropentanoic acid (AFPA), which are irreversible, mechanism-based inhibitors of pyridoxal phosphatedependent enzymes. Spontaneous mutants of Synechococcus PCC6301 resistant to these inhibitors contain altered enzyme that displays corresponding resistance to high concentrations of the inhibitor. The enzyme from strain GR6, resistant to both inhibitors, contains a three-amino-acid deletion at positions 5–7 and a Met248 → Ile substitution. The enzyme from strain K40 resistant to AFPA but not to gabaculine, contains a Ser163 → Thr substitution. GSA aminotransferases containing either the deletion or the substitution that are characteristic of the GR6 mutant were produced in Escherichia coli using the expression vector pMalc2. These engineered mutant enzymes were characterized in terms of their catalytic parameters and sensitivities to gabaculine and AFPA. Furthermore, maltose binding protein/aminotransferase fusion proteins were characterized spectrophotometrically to monitor the interaction of bound cofactor with diamino- and dioxocompounds related to the substrate and both inhibitors. Results were compared with those for similarly produced recombinant wild-type, K40 and GR6 GSA aminotransferases. The engineered products with either the N-terminal deletion or the Met248 → Ile substitution displayed catalytic efficiencies that were intermediate between the wild-type and GR6 or K40 enzymes. However, with respect to their absorption spectra, sensitivity to inhibitors and the reactivity of bound cofactor, they were essentially wild-type. These in vitro studies demonstrate that both changes in enzyme structure are necessary to obtain the distinctive properties of the GR6 aminotransferase, including resistance to high concentrations of gabaculine and AFPA. 相似文献
5.
Angelica Giancaspro Daniele Rosellini Antonio Blanco Agata Gadaleta 《Plant Cell, Tissue and Organ Culture》2012,109(3):447-455
Selectable marker genes are widely used for the efficient transformation of crop plants. In most cases, selection is based on antibiotic or herbicide resistance genes because they tend to be most efficient. The Synechococcus hemL gene has been successfully employed as a selectable marker for tobacco and alfalfa genetic transformation, by using gabaculine as the selective agent. The gene conferring gabaculine resistance is a mutant form of the hemL gene from Synechococcus PCC6301, strain GR6, encoding a gabaculine insensitive form of the glutamate1-semialdehyde aminotransferase (GSA) enzyme. In the present study we compared the transformation and selection efficiency of the common selection method based on the Streptomyces hygroscopicus bar gene conferring resistance to Bialaphos®, with both the Synechococcus hemL gene and a Medicago sativa mutated GSA gene (MsGSAgr) conferring resistance to phytotoxin gabaculine. Callus derived from immature embryos of the durum wheat cultivar Varano were simultaneously co-bombarded with bar/hemL and bar/MsGSAgr genes. After gene delivery, the marker genes were individually evaluated through all the selection phases from callus regeneration to adult plant formation, and compared for their transformation and selection efficiency. The integration of the three genes in the T0 generation was confirmed by PCR analysis with specific primers for each gene and southern blot analysis. Both Synechococcus hemL and MsGSA were more efficient than bar for biolistic transformation (2.8% vs. 1.8% and 1.1% vs. 0.5%) and selection (79% vs. 43% and 87% vs. 50%). Thus, an efficient selection method for durum wheat transformation was established that obviates the use of herbicide resistance genes. 相似文献
6.
The biodegradation of tributyl phosphate (Bu3-P, TBP), releasing phosphate at a high enough concentration locally to precipitate uranium from solution, was demonstrated
by a mixed culture consisting primarily of pseudomonads. The effect of various parameters on Bu3-P biodegradation by growing cells is described. Growth at the expense of Bu3-P as the carbon and phosphorus source occurred over a pH range from 6.5 to 8, and optimally at pH 7. Bu3-P biodegradation was optimal at 30 °C, reduced at 20 °C and negligible at 4 °C and 37 °C. Incorporation of Cu or Cd inhibited,
and Ni, Co and Mn reduced its degradation. Inorganic phosphate (above 10 mM) and kerosene (up to 1 g/l) reduced Bu3-P biodegradation significantly, but nitrate had no effect. Sulphate (10–100 mM) was inhibitory. When pregrown biomass was used
the fastest rates of tributyl and dibutyl phosphate biodegradation were 25 μmol h−1 mg protein−1 and 37 μmol h−1 mg protein−1 respectively. Microcarrier-immobilised biomass decontaminated uranium-bearing acid mine waste water by uranium phosphate
precipitation at the expense of Bu3-P hydrolysis in the presence of 35 mM SO4
2−. At pH 4.5, 79% of the UO2
2+ was removed at a flow rate of 1.4 ml/h on a 7-ml test column.
Received: 2 June 1997 / Received revision: 15 September 1997 / Accepted: 19 September 1997 相似文献
7.
Cyanobacteria were a major constituent of phototrophic communities in the lakes, ponds and streams of Bylot Island, in the
Canadian high Arctic. The waters spanned a range of temperatures (1.8–16.8°C in late July), pH regimes (6.2–9.2) and conductivities
(1.5–1700 μS cm−1) but nutrient concentrations were consistently low (< 1 μg dissolved reactive P l−1 at all sites; < 10 μg NO3-N l−1 at most sites). Picoplanktonic species (Synechococcus spp.) were often the numerical dominants in the plankton, and periphytic filamentous species (Oscillatoriaceae) commonly
formed thick (5–50 mm) benthic mats. Bloom-forming species of cyanobacteria were either absent or poorly represented even
in Chla-rich ponds. The total community biomass ranged from 0.1 to 29.8 μg Chla l−1 in the plankton and from 1.1 to 34.8 μg Chla cm−2 in the benthos. The in vivo absorbance characteristics of isolates from these environments indicated a genetically diverse
range of species in each group of Arctic cyanobacteria. Growth versus irradiance relationships were determined for each of
the isolates and similarly revealed large genetic differences (maximum growth rates from 0.17 to 0.61 day−1), even between morphologically identical taxa. A comparison of nutrients, pigment concentrations and species composition
underscores the strong similarities between freshwater ecosystems in the north and south polar zones.
Received: 3 June 1996 / Accepted: 3 November 1996 相似文献
8.
Ahlert Schmidt 《Archives of microbiology》1980,127(3):259-265
Two different thioredoxins designated as thioredoxin A and B have been isolated from the cyanobacterium Synechococcus 6301. Methods for large scale purification of these thioredoxins were developed. Thioredoxin B has been purified to homogeneity; it has a molecular weight of 11,800 and an isoelectric point of 4.6. The following K
m data were obtained for this thioredoxin; a) in the PAPS-sulfotransferase assay of Synechococcus 6301: 10.7 M; b) in the fructose-1-6-bisphosphatase assay of Synechococcus 6301: 1.7 M; c) in the APS-sulfotransferase assay of Chroococcidiopsis 7203: 5.4M. Thioredoxin A has an isoelectric point of 4.1 and it is active in the PAPS-sulfotransferase and fructose-1-6-bisphosphatase of Synechococcus 6301; it is not active in the APS-sulfotransferase of Chroococcidiopsis 7203.Dedicated to Professor Dr. O. Kandler on the occasion of his 60th birthday 相似文献
9.
A mixed culture of microorganisms able to utilize 4,6-dinitro-ortho-cresol (DNOC) as the sole source of carbon, nitrogen and energy was isolated from soil contaminated with pesticides and from
activated sludge. DNOC was decomposed aerobically in batch cultures as well as in fixed-bed column reactors. Between 65% and
84% of the substrate nitrogen was released as nitrate into the medium, and 61% of the carbon from uniformly 14C-labelled DNOC was recovered as 14CO2. The mixed microbial culture also decomposed 4-nitrophenol and 2,4-dinitrophenol but not 2,3-dinitrophenol, 2,6-dinitrophenol,
2,4-dinitrotoluene, 2,4-dinitrobenzoic acid or 2-sec-butyl-4,6-dinitrophenol (Dinoseb). Maximal degradation rates for DNOC by the bacterial biofilm immobilized on glass beads
in fixed-bed column reactors were 30 mmol day−1 (l reactor volume)−1, leaving an effluent concentration of less than 5 μg l−1 DNOC in the outflowing medium. The apparent K
s value of the immobilized mixed culture for DNOC was 17 μM. Degradation was inhibited at DNOC concentrations above 30 μM and
it ceased at 340 μM, possibly because of the uncoupling action of the nitroaromatic compound on the cellular energy-transducing
mechanism.
Received: 27 March 1997 / Received revision: 5 June 1997 / Accepted: 7 June 1997 相似文献
10.
We investigated seasonal variation of grazing impact of the pigmented nanoflagellates (PNF) with different sizes upon Synechococcus in the subtropical western Pacific coastal waters using grazing experiments with fluorescently labeled Synechococcus (FLS). For total PNF, conspicuous seasonal variations of ingestion rates on Synechococcus were found, and a functional response was observed. To further investigate the impact of different size groups, we separated
the PNF into four categories (<3, 3–5, 5–10, and >10 μm). Our results indicated that the smallest PNF (<3 μm PNF) did not
ingest FLS and was considered autotrophic. PNF of 3–5 μm in size made up most of the PNF community; however, their ingestion
on Synechococcus was too low (0.1–1.9 Syn PNF−1 h−1) to support their growth, and they had to depend on other prey or photosynthesis to survive. The ingestion rate of the 3–5 μm
group exhibited no significant seasonal variation; by contrast, the ingestion rates of 5–10 and >10 μm PNFs showed significant
seasonal variation. During the warm season, 3–5 μm PNF were responsible for the grazing of 12% of Synechococcus production, 5–10 μm PNF for 48%, and >10 μm PNF for 2%. Taken together, our results demonstrate that the PNF of 3–10 μm consumed
most Synechococcus during the warm season and exhibited a significant functional response to the increase in prey concentration. 相似文献
11.
Picophytoplankton biomass, community structure and productivity in the Great Astrolabe Lagoon, Fiji 总被引:4,自引:0,他引:4
Phytoplankton biomass, community structure and productivity of the Great Astrolabe lagoon and surrounding ocean were studied
using measurements of chlorophyll concentration and carbon uptake. The contribution of picophytoplankton to biomass, productivity
and community structure was estimated by size fractionation, 14C-incubation and flow cytometry analysis. Picoplankton red fluorescence was demonstrated to be a proxy for chlorophyll <3 μm.
Consequently, the percentage contribution to chl a<3 μm from each picoplankton group could be calculated using regression estimated values of ψ
i
(fg chl a per unit of red fluorescence). In the lagoon, average chlorophyll concentration was 0.8 mg m-3 with 45% of phytoplankton <3 μm. Primary production reached 1.3 g C m-2 day-1 with 53% due to phytoplankton <3 μm. Synechococcus was the most abundant group at all stations, followed by Prochlorococcus and picoeukaryotes. At all stations, Prochlorococcus represented less than 4% of the chl a <3 μm, Synechococcus between 85 and 95%, and Picoeukaryotes between 5 and 10%. In the upper 40 m of surrounding oceanic waters, phytoplankton
biomass was dominated by the >3 μm size fraction. In deeper water, the <1 μm size fraction dominated. Prochlorococcus was the most abundant picoplankton group and their contributions to the chlorophyll a<3 μm were close to that of the picoeukaryotes (50% each).
Accepted: 27 May 1999 相似文献
12.
Klaus-Peter Michel Pablo Exss-Sonne Gabriele Scholten-Beck Uwe Kahmann Hans Georg Ruppel Elfriede K. Pistorius 《Planta》1998,205(1):73-81
Iron-deficiency-induced protein A (IdiA) with a calculated molecular mass of 35 kDa has previously been shown to be essential
under manganese- and iron-limiting conditions in the cyanobacteria Synechococcus PCC 6301 and PCC 7942. Studies of mutants indicated that in the absence of IdiA mainly photosystem II becomes damaged, suggesting
that the major function of IdiA is in Mn and not Fe metabolism (Michel et al. 1996, Microbiology 142: 2635–2645). To further elucidate the function of IdiA, the immunocytochemical localization of IdiA
in the cell was examined. These investigations provided evidence that under mild Fe deficiency IdiA is intracellularly localized
and is mainly associated with the thylakoid membrane in Synechococcus PCC 6301. The protein became distributed throughout the cell under severe Fe limitation when substantial morphological changes
had already occurred. For additional verification of a preferential thylakoid membrane association of IdiA, these investigations
were extended to the thermophilic Synechococcus elongatus. In this cyanobacterium Mn deficiency could be obtained more rapidly than in the mesophilic Synechococcus PCC 6301 and PCC 7942, and the thylakoid membrane structure proved to be more stable under limiting growth conditions. The
immunocytochemical investigations with this cyanobacterium clearly supported a thylakoid membrane association of IdiA. In
addition, evidence was obtained for a localization of IdiA on the cytoplasmic side of the thylakoid membrane. All available
data support a function of IdiA as an Mn-binding protein that facilitates transport of Mn via the thylakoid membrane into
the lumen to provide photosystem II with Mn. A possible explanation for the observation that IdiA was not only expressed under
Mn deficiency but also under Fe deficiency is given in the discussion.
Received: 28 July 1997 / Accepted: 26 November 1997 相似文献
13.
Oze H Hirao M Ebina K Shi K Kawato Y Kaneshiro S Yoshikawa H Hashimoto J 《In vitro cellular & developmental biology. Animal》2012,48(2):123-130
Previous studies have demonstrated that oxygen environment is an important determinate factor of cell phenotypes and differentiation,
although factors which affect pericellular oxygen concentration (POC) in murine chondrogenic cell culture remain unidentified.
Oxygen concentrations in vivo were measured in rabbit musculoskeletal tissues, which were by far hypoxic compared to 20% O2 (ranging from 2.29 ± 1.16 to 4.36 ± 0.51%). Oxygen concentrations in murine chondrogenic cell (C3H10T1/2) culture medium
were monitored in different oxygen concentrations (20% or 5%) in the incubator and in different medium volumes (3,700 or 7,400 μl)
within 25-cm2 flasks. Chondrogenic differentiation was assessed by glycosaminoglycan production with quantitative evaluation of Alcian
blue staining in 12-well culture dishes. Expression of chondrogenic genes, aggrecan, and type II collagen α1, was examined
by quantitative real-time polymerase chain reaction. Oxygen concentrations in medium decreased accordingly with the depth
from medium surface, and POC at Day 6 was 18.99 ± 0.81% in 3,700-μl medium (1,480-μm depth) and 13.26 ± 0.23% in 7,400-μl
medium (2,960-μm depth) at 20% O2 in the incubator, which was 4.96 ± 0.08% (1,480-μm depth) and 2.83 ± 0.42% (2,960-μm depth) at 5% O2, respectively. The differences of POC compared by medium volume were statistically significant (p = 0.0003 at 20% and p = 0.001 at 5%). Glycosaminoglycan production and aggrecan gene expression were most promoted when cultured in moderately
low POC, 1,000 μl (2,960-μm depth) at 20% O2 and 500 μl (1,480-μm depth) at 5% O2 in 12-well culture dishes. We demonstrate that medium volume and oxygen concentration in the incubator affect not only POC
but also chondrogenic differentiation. 相似文献
14.
Inhibition of electron transport activities in the spheroplasts ofSynechococcus 6301 by HgCl2 is dependent on the concentration of mercury ions. The inhibition of whole chain electron transport activity occurs at low
concentration of Hg2+ (6 ΜM@#@). This inhibition occurs mostly due to interaction of Hg2+ on plastocyanin. At an elevated concentration (24 ΜM@#@), mercury induces inhibition chiefly in photosystem II catalyzed electron transport. At this concentration it also alters
both the absorption and emission characteristics of the phycocyanin. The photosystem I catalyzed electron transport was inhibited
by 50% only at high concentrations (36 ΜM@#@) of HgCl2. However, electron transport catalyzed by photosystems I and II from reduced duroquinone to methylviologen which involves
intersystem electron transport is extremely sensitive to mercury (low concentration 6–9 ΜM) like that of whole chain assay
indicating that the observed inhibition in whole chain electron transport at low concentrations is mostly contributed by the
damage involving other intersystem electron transport carrier(s) like plastocyanin. Thus mercury ions depending on the concentration
affects the electron transport at multiple sites in the spheroplasts ofSynechococcus. 相似文献
15.
Malaysia is the world’s leading producer of palm oil products that contribute US$ 7.5 billion in export revenues. Like any
other agro-based industries, it generates waste that could be utilized as a source of organic nutrients for microalgae culture.
Present investigation delves upon Isochrysis sp. culture in POME modified medium and its utilization as a supplement to Nanochloropsis sp. in rotifer cultures. The culture conditions were optimized using a 1 L photobioreactor (Temp: 23°C, illumination: 180 ∼ 200 μmol
photons m−2s−1, n = 6) and scaled up to 10 L outdoor system (Temp: 26–29°C, illumination: 50 ∼ 180 μmol photons m−2s−1, n = 3). Algal growth rate in photobioreactor (μ = 0.0363 h−1) was 55% higher compared to outdoor culture (μ = 0.0163 h−1), but biomass production was 1.3 times higher in outdoor culture (Outdoor = 91.7 mg m−2d−1; Photobioreactor = 69 mg m−2d−1). Outdoor culture produced 18% higher lipid; while total fatty acids (FA) was not significantly affected by the change in
culture systems as both cultures yield almost similar concentrations of fatty acids per gram of sample (photobioreactor = 119.17 mg
g−1; outdoor culture = 104.50 mg g−1); however, outdoor cultured Isochrysis sp. had 26% more polyunsaturated fatty acids (PUFAs). Rotifers cultured in Isochrysis sp./ Nanochloropsis sp. (1:1, v/v) mixture gave similar growth rate as 100% Nanochoropsis sp. culture (μ = 0.40 d−1), but had 45% higher counts of rotifers with eggs (t = 7, maximum). The Isochrysis sp. culture successfully lowered the nitrate (46%) and orthophosphate (83%) during outdoor culture. 相似文献
16.
Thomas Rohde Sven Asp Dave A. MacLean Bente K. Pedersen 《European journal of applied physiology and occupational physiology》1998,78(5):448-453
This study examined whether oral glutamine supplementation abolishes some of the exercise-induced changes in lymphocyte functions
following long-term intense exercise. A group of 16 marathon runners participating in The Copenhagen Marathon 1996 were placed
randomly in either a placebo (n = 7) or a glutamine receiving group (n = 9). Each subject received four doses of either placebo or glutamine (100 mg · kg−1) administered at 0, 30, 60, and 90-min post-race. In the placebo group the plasma glutamine concentrations were lower than
pre-race values during the post-exercise period [mean 647 (SEM 32) compared to 470 (SEM 22) μmol · l−1 90-min post-race, P < 0.05] whereas glutamine supplementation maintained the plasma glutamine concentration (at ∼750 μmol · l−1). Glutamine supplementation in vivo had no effect on the lymphokine activated killer (LAK) cell activity, the proliferative
responses or the exercise-induced changes in concentrations or percentages of any of the leucocyte subpopulations examined.
Glutamine addition in in vitro studies enhanced the proliferative response in both groups. These data would suggest that decreased
plasma glutamine concentrations post-exercise are not responsible for exercise-induced decrease in LAK activity and that the
influence of glutamine in vitro is not dependent on the plasma glutamine concentration at the time of sampling.
Accepted: 23 April 1998 相似文献
17.
Potential uptake and clearance rates of fluorescent microspheres (FM) from 0.25 to 4.05 μm diameter were determined for the
non-loricate ciliate Pseudocohnilembus sp. from Antarctic sea ice. The percentage of ciliate cells that ingested FM after 20 min incubation decreased with increasing
particle diameter. Pseudocohnilembus sp. ingested FM between 0.25 and 4.05 μm in diameter. We offered FM at concentrations less than natural concentrations for
plankton plus detrital material and obtained clearance rates less than those previously reported for bactivorous ciliates.
Clearance rates were 3.6–5.4 nl cell−1 h−1 for FM 0.5 and 1 μm diameter, respectively, but decreased to 1.1 nl cell−1 h−1 for 1.97 μm diameter and 1.4 nl cell−1 h−1 for 4.05-μm-diameter FM. Clearance and uptake rates of FM 0.5 and 1 μm diameter indicate that Pseudocohnilembus sp. principally grazes on bacteria-sized particles. However, it can also ingest organisms as large as nanoplankton and may
graze particles as small as femtoplankton and colloids. This suggests a feeding strategy that may suit the temporal and spatial
changes in food availability in the sea-ice habitat.
Accepted: 13 August 2000 相似文献
18.
19.
An DS Cui CH Sung BH Yang HC Kim SC Lee ST Im WT Kim SG 《Applied microbiology and biotechnology》2012,94(3):673-682
The gene encoding an α-l-arabinofuranosidase that could biotransform ginsenoside Rc {3-O-[β-d-glucopyranosyl-(1–2)-β-d-glucopyranosyl]-20-O-[α-l-arabinofuranosyl-(1–6)-β-d-glucopyranosyl]-20(S)-protopanaxadiol} to ginsenoside Rd {3-O-[β-d-glucopyranosyl-(1–2)-β-d-glucopyranosyl]-20-O-β-d-glucopyranosyl-20(S)-protopanaxadiol} was cloned from a soil bacterium, Rhodanobacter ginsenosidimutans strain Gsoil 3054T, and the recombinant enzyme was characterized. The enzyme (AbfA) hydrolyzed the arabinofuranosyl moiety from ginsenoside
Rc and was classified as a family 51 glycoside hydrolase based on amino acid sequence analysis. Recombinant AbfA expressed
in Escherichia coli hydrolyzed non-reducing arabinofuranoside moieties with apparent K
m values of 0.53 ± 0.07 and 0.30 ± 0.07 mM and V
max values of 27.1 ± 1.7 and 49.6 ± 4.1 μmol min−1 mg−1 of protein for p-nitrophenyl-α-l-arabinofuranoside and ginsenoside Rc, respectively. The enzyme exhibited preferential substrate specificity of the exo-type
mode of action towards polyarabinosides or oligoarabinosides. AbfA demonstrated substrate-specific activity for the bioconversion
of ginsenosides, as it hydrolyzed only arabinofuranoside moieties from ginsenoside Rc and its derivatives, and not other sugar
groups. These results are the first report of a glycoside hydrolase family 51 α-l-arabinofuranosidase that can transform ginsenoside Rc to Rd. 相似文献
20.
Vishal Gupta Manoj Kumar Puja Kumari C. R. K. Reddy Bhavanath Jha 《Journal of applied phycology》2011,23(2):209-218
This study reports on the optimization of protoplast yield from two important tropical agarophytes Gracilaria dura and Gracilaria verrucosa using different cell-wall-degrading enzymes obtained from commercial sources. The conditions for achieving the highest protoplast
yield was investigated by optimizing key parameters such as enzyme combinations and their concentrations, duration of enzyme
treatment, enzyme pH, mannitol concentration, and temperature. The significance of each key parameter was also further validated
using the statistical central composite design. The enzyme composition with 4% cellulase Onozuka R-10, 2% macerozyme R-10,
0.5% pectolyase, and 100 U agarase, 0.4 M mannitol in seawater (30‰) adjusted to pH 7.5 produced the highest protoplast yields
of 3.7 ± 0.7 × 106 cells g−1 fresh wt for G. dura and 1.2 ± 0.78 × 106 cells g−1 fresh wt for G. verrucosa when incubated at 25°C for 4–6 h duration. The young growing tips maximally released the protoplasts having a size of 7–15 μm
in G. dura and 15–25 μm in G. verrucosa, mostly from epidermal and upper cortical regions. A few large-size protoplasts of 25–35 μm, presumably from cortical region,
were also observed in G. verrucosa. 相似文献