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1.
1H-(13)C one-bond dipolar coupling values were measured for natural abundance samples of the human milk oligosaccharides "lacto-N-fucopentaose" (LNF-1 LNF-2, and LNF-3), "lacto-N-difucohexaose" (LND-1), "lacto-N-tetraose" (LNT), and "lacto-N-neo-tetraose" (LNnT), four of which have Lewis blood group epitopes. Each oligosaccharide was dissolved in a 7.5% solution of 1, 2-dimyristoyl-sn-glycero-3-phosphocholine/1, 2-dihexanoyl-sn-glycero-3-phosphocholine (DMPC/DHPC) bicelle liquid crystals oriented in the NMR magnetic field. The dipolar coupling data and NOE were fitted to conformational models with calculations of an optimum orientation tensor which best represents the dipolar coupling values for a fragment hypothesized to adopt a single conformation. In the case of LNF-1, LNF-2, LNF-3, and LND-1, the models confirm previous conformational models for the Lewis epitopes based on NOE and molecular dynamics simulations. Extensions of the model provided new structural data for the remaining residues. In all cases, upper limits for the errors in the glycosidic angles of the models were estimated. Since residual dipolar coupling provides information on long-range order, it is a valuable complement to other types of NMR data such as NOE and scalar coupling for exploring conformations of complex oligosaccharides. 相似文献
2.
Through control of both the nmr probe temperature and of the solvent viscosity, phase-sensitive two-dimensional 1H nuclear Overhauser data (NOESY) at 300 and 500 MHz are obtained with excellent signal-to-noise ratios for Lewis blood group penta- and hexasaccharides isolated from human milk. Relatively long mixing times are required to produce measurable NOE intensities in these oligosaccharides, which makes a full relaxation matrix analysis necessary. By measurements of selective T1 for a few isolated 1H resonances, it was possible to generate a simulation of the complete NOESY spectrum at arbitrary mixing time for comparison with the experimental data. From an exhaustive search of the conformational space, it was found that only a small range of glycosidic dihedral angles of the nonreducing terminal Lewis blood group determinant fragments of the milk oligosaccharides LNF-2 and LND-1 produce simulated spectra agreeing within experimental error to the data. Conformational energy calculations reveal that each of these conformations is also one of minimum energy. It is concluded that the Lewis(a) and Lewis(b) oligosaccharides adopt relatively compact rigid structures in solution, as shown by the observation of cross peaks between protons in nonadjacent residues. Like the blood group A and H oligosaccharides, there exists only a small dependence of the conformation for Lewis(a) and Lewis(b) oligosaccharides on solvent. The apparent lack of dependence of conformation of these oligosaccharides on DMSO in D2O suggests that modification of solvent viscosity with mixtures of DMSO:D2O may provide a useful general strategy of NOESY studies of oligosaccharides. 相似文献
3.
Sakamoto T Tanaka H Nishimura Y Ishimaru M Kasai N 《Applied microbiology and biotechnology》2011,90(5):1701-1710
A type II arabinogalactan-degrading enzyme, termed Exo-1,3-Gal, was purified to homogeneity from the culture filtrate of Sphingomonas sp. 24T. It has an apparent molecular mass of 48 kDa by SDS–PAGE. Exo-1,3-Gal was stable from pH 3 to 10 and at temperatures
up to 40 °C. The optimum pH and temperature for enzyme activity were pH 6 to 7 and 50 °C, respectively. Galactose was released
from β-1,3-d-galactan and β-1,3-d-galactooligosaccharides by the action of Exo-1,3-Gal, indicating that the enzyme was an exo-β-1,3-d-galactanase. Analysis of the reaction products of β-1,3-galactotriose by high-performance anion-exchange chromatography revealed
that the enzyme hydrolyzed the substrate in a non-processive mode. Exo-1,3-Gal bypassed the branching points of β-1,3-galactan
backbones in larch wood arabinogalactan (LWAG) to produce mainly galactose, β-1,6-galactobiose, and unidentified oligosaccharides
1 and 2 with the molar ratios of 7:19:62:12. Oligosaccharides 1 and 2 were enzymatically determined to be β-1,6-galactotriose
and β-1,6-galactotriose substituted with a single arabinofuranose residue, respectively. The ratio of side chains enzymatically
released from LWAG was in good agreement with the postulated structure of the polysaccharide previously determined by chemical
methods. 相似文献
4.
Syntheses of l-dopa 1a glucoside 10a,b and dl-dopa 1b glycosides 10–18 with d-glucose 2, d-galactose 3, d-mannose 4, d-fructose 5, d-arabinose 6, lactose 7, d-sorbitol 8 and d-mannitol 9 were carried out using amyloglucosidase from Rhizopus mold, β-glucosidase isolated from sweet almond and immobilized β-glucosidase. Invariably, l-dopa and dl-dopa gave low to good yields of glycosides 10–18 at 12–49% range and only mono glycosylated products were detected through glycosylation/arylation at the third or fourth
OH positions of l-dopa 1a and dl-dopa 1b. Amyloglucosidase showed selectivity with d-mannose 4 to give 4-O-C1β and d-sorbitol 8 to give 4-O-C6-O-arylated product. β-Glucosidase exhibited selectivity with d-mannose 4 to give 4-O-C1β and lactose 7 to give 4-O-C1β product. Immobilized β-glucosidase did not show any selectivity. Antioxidant and angiotensin converting enzyme inhibition
(ACE) activities of the glycosides were evaluated glycosides, out of which l-3-hydroxy-4-O-(β-d-galactopyranosyl-(1′→4)β-d-glucopyranosyl) phenylalanine 16 at 0.9 ± 0.05 mM and dl-3-hydroxy-4-O-(β-d-glucopyranosyl) phenylalanine 11b,c at 0.98 ± 0.05 mM showed the best IC50 values for antioxidant activity and dl-3-hydroxy-4-O-(6-d-sorbitol)phenylalanine 17 at 0.56 ± 0.03 mM, l-dopa-d-glucoside 10a,b at 1.1 ± 0.06 mM and dl-3-hydroxy-4-O-(d-glucopyranosyl)phenylalanine 11a-d at 1.2 ± 0.06 mM exhibited the best IC50 values for ACE inhibition.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
5.
Purification and functional characterization of endo-β-mannanase MAN5 and its application in oligosaccharide production from konjac flour 总被引:1,自引:0,他引:1
Min Zhang Xiu-Lan Chen Zhi-Hua Zhang Cai-Yun Sun Lei-Lei Chen Hai-Lun He Bai-Cheng Zhou Yu-Zhong Zhang 《Applied microbiology and biotechnology》2009,83(5):865-873
MAN5, the main extracellular saccharide hydrolase from Bacillus sp. MSJ-5, is an endo-β-mannanase with a demand of at least five sugar moieties for effective cleavage. It has a pH optimum
of 5.5 and a temperature optimum of 50°C and is stable at pH 5–9 or below 65°C. MAN5 has a very high ability to hydrolyze
konjac flour, 10 U/mg of which could completely liquefy konjac flour gum in 10 min at 50°C. HPLC analysis showed that most
glucomannan in the konjac flour was hydrolyzed into a large amount of oligosaccharides with DP of 2–6 and a very small amount
of monosaccharide. With the culture supernatant as enzyme source, the optimum condition to prepare oligosaccharides from konjac
flour was obtained as 10 mg/ml konjac flour incubated with 10 U/mg enzyme at 50°C for 24 h. With this condition, more than
90% polysaccharides in the konjac flour solution were hydrolyzed into oligosaccharides and a little monosaccharide (2.98%
of the oligosaccharides). Konjac flour is an underutilized agricultural material with low commercial value in China. With
MAN5, konjac flour can be utilized to generate high value-added oligosaccharides. The high effectiveness and cheapness of
this technique indicates its potential in industry.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.
Min Zhang and Xiu-Lan Chen contributed equally to this work. 相似文献
6.
7.
Despina K. Laimou Maria Katsara Minos-Timotheos I. Matsoukas Vasso Apostolopoulos Anastassios N. Troganis Theodore V. Tselios 《Amino acids》2010,39(5):1147-1160
Leuprolide [dLeu6, NHEt10]GnRH, a potent gonadotropin-releasing hormone (GnRH) agonist, is used in a wide variety of hormone-related diseases like
cancer and endometriosis. In this report, the conformational behaviour of Leuprolide and its linear synthetic analogues, namely
[Tyr5(OMe), dLeu6, Aze9, NHEt10]GnRH (1) and [Tyr5(OMe), dLeu6, NHEt10]GnRH (2) have been studied in DMSO and H2O solutions by means of 2D nuclear magnetic resonance (NMR) experiments and detailed molecular dynamics (MD) simulations.
The aim was to identify the conformational requirements of GnRH analogues for agonistic activity. This approach is of value
as no crystallographic data are available for the GnRH receptor (G protein-coupled receptor, GPCR). The NOE data indicate
the existence of a β-turn type I in the 2–5 segments of Leuprolide and its linear analogues in the case of using DMSO-d6 as solvent, whereas a β-turn type II in the 3–6 segments is indicated using D2O as solvent. The final structures fulfil the conformational requirements that are known, in the literature, to play a significant
role in receptor recognition and activation. Finally, the linear analogues (1) and (2) are biologically active when tested against the human breast cancer cell line, MCF-7. 相似文献
8.
Paula Alayón-Luaces Eduardo A. Pagano Luis A. Mroginski Gabriel O. Sozzi 《Plant Cell, Tissue and Organ Culture》2010,101(1):1-10
Sucrose presence and concentration modulated in different ways and to different extents the activity of six plant glycoside
hydrolases (PGHs) extracted from apple callus cultures, both in the water soluble fraction (WS-F) and in the NaCl-released
fraction (NaCl-F). β-d-Glucosidase activity increased because of sucrose starvation and the addition of sucrose decreased both WS-F and NaCl-F β-d-glucosidase from calli grown in a Murashige and Skoog’s basal medium with (MSH) or without (MS0) plant growth regulators
(PGRs). WS-F and NaCl-F α-l-arabinofuranosidase, NaCl-F β-d-galactosidase and NaCl-F β-d-xylosidase activity reached a maximum when 0.045 M sucrose was added to the MS0 medium with an ensuing decline at higher
sucrose concentrations. α-d-Galactosidase and α-d-xylosidase activity reached a maximum when 0.045 M sucrose was supplied and did not decline significantly in 0.09 M sucrose-supplied
calli. When the effects of PGR presence or absence were analysed, NaCl-F β-d-glucosidase, α-d-galactosidase, β-d-galactosidase, α-d-xylosidase and β-d-xylosidase activities were found to be higher in MS0 than in MSH. To assess whether sugar effects were sucrose-specific,
other sugars (glucose, fructose, galactose, maltose, lactose, raffinose, sorbitol and mannitol) were tested, with or without
PGR supplementation. In general, sugar alcohols (mannitol, sorbitol) and some monosaccharides (fructose and glucose in particular)
were better inducers of NaCl-F α-l-arabinofuranosidase, β-d-galactosidase and β-d-xylosidase activity than disaccharides (sucrose, maltose, and lactose) or the trisaccharide raffinose. This trend was not
widespread to all PGHs assessed since sucrose-supplemented calli displayed higher NaCl-F α-d-galactosidase than those supplemented with glucose, galactose, sorbitol or mannitol. These results show that sugars supplied
to callus tissue cultures as a carbon source can also modulate PGH activity. Modulation is different for each PGH, sugar-specific
and, at least in the case of sucrose, concentration-dependent. Results also suggest the existence of regulatory interactions
between PGRs and sugars as part of an intricate sensing and signalling network. Combination of PGR, sugar type and concentration
should be taken into account to maximize each PGH activity for further enzyme studies. 相似文献
9.
Accumulating evidence suggests that Aβ1–42–membrane interactions may play an important role in the pathogenesis of Alzheimer’s disease. However, the mechanism of this
structural transition remains unknown. In this work, we have shown that submicellar concentrations of sodium dodecyl sulfate
(SDS) can provide a minimal platform for Aβ1–42 self-assembly. To further investigate the relation between Aβ1–42 structure and function, we analyzed peptide conformation and aggregation at various SDS concentrations using circular dichroism
(CD), Fourier transform infrared spectroscopy, and gel electrophoresis. These aggregates, as observed via atomic force microscopy,
appeared as globular particles in submicellar SDS with diameters of 35–60 nm. Upon sonication, these particles increased in
disc diameter to 100 nm. Pyrene I
3/I
1 ratios and 1-anilinonaphthalene-8-sulfonic acid binding studies indicated that the peptide interior is more hydrophobic than
the SDS micelle interior. We have also used Forster resonance energy transfer between N-terminal labeled pyrene and tyrosine
(10) of Aβ1–42 in various SDS concentrations for conformational analysis. The results demonstrate that SDS at submicellar concentrations
accelerates the formation of spherical aggregates, which act as niduses to form large spherical aggregates upon sonication.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
10.
V. Jaitak V. Kumar Kaul Bandna N. Kumar B. Singh L. S. Savergave V. V. Jogdand S. Nene 《Biotechnology letters》2009,31(9):1415-1420
Stevioside was subjected to 1,4-intermolecular transglycosylation using β-cyclodextrin glucanotransferase (β-CGtase) produced
from an alkalophilic strain of Bacillus firmus. The reaction was carried out by traditional, ultrasound-assisted and microwave-assisted techniques. Reaction under microwave
conditions was faster and was completed in 1 min yielding two 1,4 transglycosylated products, 4′-O-alpha-d-glycosyl stevioside (I) and 4′′-O-alpha-d-maltosyl stevioside (II) in 66% and 24%, respectively. The optimum transglycosylation occurred by using stevioside (1.24 mmol),
β-CD (1.76 mmol) and β-CGtase (2 U/g) under microwave assisted reaction (MAR) in 5 ml sodium phosphate buffer (pH 7) at 50°C
and 80 W power. MAR is therefore potentially a useful and economical method for faster transglycosylation of stevioside.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
11.
The use of NMR residual dipolar couplings in aqueous dilute liquid crystalline medium for conformational studies of complex oligosaccharides 总被引:1,自引:0,他引:1
C-H dipolar coupling values were measured for a natural-abundance sample of the pentasaccharide beta-D-Galp-(1-->3)-[alpha-L-Fucp-(1-->4)]-beta-D-GlcNAcp-(1 -->3)-beta-D- Galp-(1-->4)-beta-D-Glcp ('lacto-N-fucopentaose 2') (LNF-2), in a 7.5% solution of dimyristoyl phosphatidylcholine-dihexanoyl phosphatidylcholine bicelle liquid crystals oriented in the NMR magnetic field. Interpretation of the dipolar coupling data and NOE confirms the conformational model for the Lewis(a) trisaccharide epitope based on NOE, molecular dynamics simulations, and scalar coupling data and provided new structural information for the remaining residues of the pentasaccharide. Since residual dipolar coupling provides information on long-range order, it is a valuable complement to other types of NMR data such as NOE and scalar coupling for exploring conformations of complex oligosaccharides. 相似文献
12.
Polyakova O Dear D Stern I Martin S Hirst E Bawumia S Nash A Dodson G Bronstein I Bayley PM 《European biophysics journal : EBJ》2009,38(2):209-218
Formation of PrP aggregates is considered to be a characteristic event in the pathogenesis of TSE diseases, accompanied by
brain inflammation and neurodegeneration. Factors identified as contributing to aggregate formation are of interest as potential
therapeutic targets. We report that in vitro proteolysis of ovine PrP94–233 (at neutral pH and in the absence of denaturants) by the protease cathepsin S, a cellular enzyme that also shows enhanced
expression in pathogenic conditions, occurs selectively in the region 135–156. This results in an unusually efficient, concentration-dependent
conformational conversion of a large subfragment of PrP94–233 into a soluble β-structured oligomeric intermediate species, that readily forms a thioflavin-T-positive aggregate. N-terminal
sequencing of the proteolysis fragments shows the aggregating species have marked sequence similarities to truncated PrP variants
known to confer unusually severe pathogenicity when transgenically expressed in PrPo/o mice. Circular dichroism analysis shows that PrP fragments 138–233, 144–233 and 156–233 are significantly less stable than
PrP94–233. This implies an important structural contribution of the β1 sequence within the globular domain of PrP. We propose that
the removal or detachment of the β1 sequence enhances β-oligomer formation from the globular domain, leading to aggregation.
The cellular implications are that specific proteases may have an important role in the generation of membrane-bound, potentially
toxic, β-oligomeric PrP species in pre-amyloid states of prion diseases. Such species may induce cell death by lysis, and
also contribute to the transport of PrP to neuronal targets with subsequent amplification of pathogenic effects.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
13.
Aditya K. Sanki Julie Boucau Donald R. Ronning Steven J. Sucheck 《Glycoconjugate journal》2009,26(5):589-596
Antigen 85 (ag85) is a complex of acyltransferases (ag85A–C) known to play a role in the mycolation of the d-arabino-d-galactan (AG) component of the mycobacterial cell wall. In order to better understand the chemistry and substrate specificity
of ag85, a trehalose monomycolate mimic p-nitrophenyl 6-O-octanoyl-β-d-glucopyranoside (1) containing an octanoyl moiety in lieu of a mycolyl moiety was synthesized as an acyl donor. Arabinofuranoside acceptors, methyl α-d-arabinofuranoside (2), methyl β-d-arabinofuranoside (3), and methyl 2-O-β-d-arabinofuranosyl-α-d-arabinofuranoside (9) were synthesized to mimic the terminal saccharides found on the AG. The acyl transfer reaction between acyl donor 1 and acceptors 2, 3, and 9 in the presence of ag85C from Mycobacterium tuberculosis (M. tuberculosis) resulted in the formation of esters, methyl 2, 5-di-O-octanoyl-α-d-arabinofuranoside (10), methyl 5-O-octanoyl-β-d-arabinofuranoside (11), and methyl 2-O-(5-O-octanoyl-β-d-arabinofuranosyl)-5-O-octanoyl-α-d-arabinofuranoside (12) in 2 h, 2 h and 8 h, respectively. The initial velocities of the reactions were determined with a newly developed assay
for acyltransferases. As expected, the regioselectivity corresponds to mycolylation patterns found at the terminus of the
AG in M. tuberculosis. The study shows that d-arabinose-based derivatives are capable of acting as substrates for ag85C-mediated acyl-transfer and the acyl glycoside 1 can be used in lieu of TMM extracted from bacteria to study ag85-mediated acyl-transfer and inhibition leading to the better understanding of
the ag85 protein class.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
14.
Hemicellulose is a complex group of heterogeneous polymers and represents one of the major sources of renewable organic matter.
Mannan is one of the major constituent groups of hemicellulose in the wall of higher plants. It comprises linear or branched
polymers derived from sugars such as d-mannose, d-galactose, and d-glucose. The principal component of softwood hemicellulose is glucomannan. Structural studies revealed that the galactosyl
side chain hydrogen interacts to the mannan backbone intramolecularly and provides structural stability. Differences in the
distribution of d-galactosyl units along the mannan structure are found in galactomannans from different sources. Acetyl groups were identified
and distributed irregularly in glucomannan. Some of the mannosyl units of galactoglucomannan are partially substituted by
O-acetyl groups. Some unusual structures are found in the mannan family from seaweed, showing a complex system of sulfated
structure. Endohydrolases and exohydrolases are involved in the breakdown of the mannan backbone to oligosaccharides or fermentable
sugars. The main-chain mannan-degrading enzymes include β-mannanase, β-glucosidase, and β-mannosidase. Additional enzymes
such as acetyl mannan esterase and α-galactosidase are required to remove side-chain substituents that are attached at various
points on mannan, creating more sites for subsequent enzymatic hydrolysis. Mannan-degrading enzymes have found applications
in the pharmaceutical, food, feed, and pulp and paper industries. This review reports the structure of mannans and some biochemical
properties and applications of mannan-degrading enzymes. 相似文献
15.
Heitz Annie Le-Nguyen Dung Castro Bertrand Chiche Laurent 《International journal of peptide research and therapeutics》1997,4(4-6):245-249
Summary Trypsin inhibitor EETI II, possessing six cysteines engaged in three disulfide bridges, shares a common structural motif with
other proteins of different origins and functions. To understand the principles that govern folding of this largely distributed
basic scaffold, mainly composed of a small triple-stranded β-sheet, we have studied different stages in the folding of EETI
II. The conformational properties of a synthetic analogue of EETI II possessing only one native (15–27) disulfide bridge were
investigated with the combined use of1H NMR and molecular modelling. Although two native-like reverse turns were observed, formation of β-sheet could not be evidenced
in the one disulfide analogue, while the motif has been shown to be present in a folding intermediate with two native disulfide
bridges (9–21 and 15–27). These results suggest that the structural motif requires stabilisation by two disulfide bridges. 相似文献
16.
Joanna Malicka Małgorzata Groth Cezary Czaplewski Regina Kasprzykowska Adam Liwo Leszek Łankiewicz Wieslaw Wiczk 《International journal of peptide research and therapeutics》1998,5(5-6):445-447
Summary The probable conformations of two cyclic enkephalin analogs, DNS-cyclo[d-Dab-Gly-Trp-Leu] (I) and DNS-cyclo[d-Dab-Gly-Trp-d-Leu] (II) (DNS=dansyl), were determined by combining the results of NOE, vicinal coupling constant and fluorescence energy transfer
measurements with theoretical calculations. The common feature of the conformations for both peptides is the presence of a
β-turn at residues 2 and 3. 相似文献
17.
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
18.
Koseki T Mese Y Fushinobu S Masaki K Fujii T Ito K Shiono Y Murayama T Iefuji H 《Applied microbiology and biotechnology》2008,77(6):1279-1285
The glycoside hydrolase family 61 endoglucanase from Aspergillus kawachii (AkCel61) is a modular enzyme that consists of a catalytic domain and a carbohydrate-binding module belonging to family 1
(CBM1) that are connected by a Ser-Thr linker region longer than 100 amino acids. We expressed the recombinant AkCel61, wild-type
enzyme (rAkCel61), and a truncated enzyme consisting of the catalytic domain (rAkCel61ΔCBM) in Pichia pastoris and analyzed their biochemical properties. Purified rAkCel61 and rAkCel61ΔCBM migrated on sodium dodecyl sulfate-polyacrylamide
gel electrophoresis (SDS-PAGE) and were demonstrated to have apparent molecular masses of 81,000 and 34,000 Da, respectively.
After treatment with endoglycosidase H, both proteins showed an increase in mobility, thus, demonstrating estimated molecular
masses of 78,000 and 28,000 Da, respectively. Mass spectrometry analysis revealed that rAkCel61 and rAkCel61ΔCBM expressed
in P. pastoris are heterogeneous due to protein glycosylation. The rAkCel61 protein bound to crystalline cellulose but not to arabinoxylan.
The rAkCel61 and rAkCel61ΔCBM proteins produced small amounts of oligosaccharides from soluble carboxymethylcellulose. They
also exhibited a slight hydrolytic activity toward laminarin. However, they showed no detectable activity toward microcrystalline
cellulose, arabinoxylan, and pectin. Both recombinant enzymes also showed no detectable activity toward p-nitrophenyl β-d-glucoside, p-nitrophenyl β-d-cellobioside, and p-nitrophenyl β-d-cellotrioside.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献
19.
Exogenously applied ABA-β-d-glucopyranosyl ester (ABA-GE) inhibited shoot growth of alfalfa (Medicago sativa L.), cress (Lepidium sativum L.), lettuce (Lactuca sativa L.), Digitaria sanguinalis L., timothy (Pheleum pratense L.) and ryegrass (Lolium multiflorum Lam.) seedlings at concentrations greater than 0.1 μM. The growth inhibitory activity of ABA-GE on these shoots was 26–40% of
that of (+)-ABA. ABA-β-d-glucosidase activities in these seedlings were 11–31 nmol mg−1 protein min−1. These results suggests that exogenously applied ABA-GE may be absorbed by plant roots and hydrolyzed by ABA-β-d-glucosidase, and liberated free ABA may induce the growth inhibition in these plants. Thus, although ABA-GE had been thought
to be physiologically inactive ABA conjugate, ABA-GE may have important physiological functions rather than an inactive conjugated
ABA form. 相似文献
20.
Fanyan Meng Chuanlai Shen Yong He Fengqin Miao Jianqiong Zhang 《Biotechnology letters》2009,31(6):831-836
The application of the peptide-linked β2-microglobulin (β2m) strategy is limited in some cases due to the incompatibility
between the sequences of the peptides and the restriction sites of the plasmid vectors. An isocaudamer technique was adapted
to overcome this restriction. Three peptide-linked β2m genes, HBc18–27-hβ2m gene, OVA257–264-mβ2m gene and HER2/neu369–377-mβ2m gene, were inserted into the pET28a vectors with this technique. The corresponding proteins were expressed in Escherichia coli with yields of over 50 mg/l culture and purities of over 80%. This strategy facilitates the construction of peptide-linked
β2m molecules and will simplify the preparation of major histocompatibility complex-peptide complexes.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. 相似文献