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1.
The genes were cloned for the two apoprotein subunits, and ,of phycocyanin from the cyanobacterium Spirulina maxima = Arthrospiramaxima) strain F3. The - and -subunit gene-coding regionscontain 489 bp and 519 bp, respectively. The -subunit gene is upstreamfrom the -subunit gene, with a 111-bp segment separating them.Similarities between the -subunits of S. maxima and nine othercyanobacteria were between 58% and 99%, as were those between the -subunits. The maximum similarity between the - and -subunits from S. maxima was 27%. 相似文献
2.
Leila M. Lopes-Alves Lucia Mendonça-Previato Bernard Fournet Pierre Degand José O. Previato 《Glycoconjugate journal》1992,9(2):75-81
-Elimination of peptidorhamnomannans purified from yeast-like and mycelial phases ofSporothrix schenckii released neutral and acidic reduced oligosaccharides that were O linked to serine and/or threonine. Man-(1–2)Man-ol, Rha(1–3)Man(1–2)Man-ol, Rha(1–4)GlcA(1–2)Man(1–2)Man-ol, and Rha(1–4)[Rha(1–2)] GlcA(1–2)Man(1–2)Man-ol were characterized based on methylation analysis, proton magnetic resonance and fast atom bombardment mass spectrometry.Abbreviations FAB
fast atom bombardment
- GLC
gas liquid chromatography
- GlcA
d-glucopyranosyluronic acid
- Man
d-mannopyranose
- Man-ol
d-mannitol
- MS
mass spectrometry
- NMR
nuclear magnetic resonance
- Rha
l-rhamnopyranose 相似文献
3.
Six tetrasaccharide fractions were isolated from shark cartilage chondroitin sulfate D by gel filtration chromatography followed by HPLC on an amine-bound silica column after exhaustive digestion with testicular hyaluronidase. Their structures were determined unambiguously by one- and two-dimensional 500 MHz1H NMR spectroscopy in conjunction with HPLC analysis of chondroitinase AC-II digests of the tetrasaccharides. One fraction was found to contain two tetrasaccharide components. All the seven tetrasaccharides shared the common core structure GlcA1-3GalNAc1-4GlcA1-3GalNAc with various sulfation profiles. Four were disulfated comprising of two monosulfated disaccharide units GlcA1-3GalNAc(4-sulfate) and/or GlcA1-3GalNAc(6-sulfate), whereas the other three were hitherto unreported trisulfated tetrasaccharides containing a disulfated disaccharide unit GlcA(2-sulfate)1-3GalNAc(6-sulfate) and a monosulfated disaccharide unit GlcA1-3GalNAc(4-or 6-sulfate). These sulfated tetrasaccharides were demonstrated to serve as appropriate acceptor substrates for serum -N-acetylgalactosaminyltransferase, indicating their usefulness as authentic oligosaccharide substrates or probes for the glycobiology of sulfated glycosaminoglycans.Abbreviations NFU
National formulary unit
- COSY
correlation spectroscopy
- HOHAHA
homonuclear Hartmann-Hahn
- 1D or 2D
one- or two-dimensional
- IdoA
l-iduronic acid
- GlcA
d-gluco-4-enepyranosyluronic acid
- Di-0S
GlcA1-3GalNAc
- Di-4S
GlcA1-3GalNAc(4-sulfate)
- Di-4S
GlcA1-3GalNAc(4-sulfate)
- Di-6S
GlcA1-3GalNAc(6-sulfate)
- Di-6S
GlcA1-3GalNAc(6-sulfate)
- Di-diS
d
GlcA(2-sulfate)1-3GalNAc(6-sulfate)
- Di-diSE
GlcA1-3GalNAc(4, 6-disulfate)
- U
G, U, 2S, 4S, and 6S represent GlcA, GalNAc, GlcA, 2-O-sulfate, 4-O-sulfate, and 6-O-sulfate, respectively 相似文献
4.
Particulate membrane preparations have been isolated from cambial cells, and from differentiating and differentiated xylem cells of the main stem of pine trees. These preparations synthesise a 14 glucomannan from guanosine 5-diphosphate-mannose. The polysaccharide and the synthase have been characterized and the Km and Vmax for the synthase determined as 85 M and 52.9 M·min-1, respectively. The enzymic activity was inhibited by the addition of guanosine 5-diphosphate-D-glucose so that the presence of an epimerase on the particulate fraction in conjunction with the synthase probably allowed the heteropolymer to be formed with the optimal ratio of the concentrations of the nucleoside-diphosphate sugar donors. No evidence for a polyprenyl-phosphate derivative as an intermediate during the polymer synthesis was obtained. Part of the control mechanism for the deposition of the large amounts of the glucomannan during the secondary thickening of the tracheids of the vascular system is by an increase in the amount of synthase activity at the endomembrane system of the cells. This probably occurs by an increase in the amount of enzyme which is modulated by gene regulation during differentiation.Abbreviations GDP
guanosine 5-diphosphate
- GLC
gasliquid chromatography 相似文献
5.
The trisaccharide 2-(p-trifluoroacetamidophenyl)ethyl 2-acetamido-2-deoxy-4-O-[2-O-(-l-fucopyranosyl)--d-galactopyranosyl]--d-glucopyranoside 1 and the tetrasaccharide 2-(p-trifluoroacetamidophenyl)ethyl 2-acetamido-2-deoxy-3-O-(-l-fucopyranosyl)-4-O-[2-O-(-l-fucopyranosyl)--d-galactopyranosyl]--d-glucopyranoside 2 were synthesized. Thioglycosides, suitably protected, activated directly with methyl trifluoromethanesulfonate or dimethyl(methylthio)sulfonium tetrafluoroborate or activated after bromine treatment with halophilic reagents, were used as glycosyl donors in the construction of the glycosidic linkages.Abbreviations DMTSB
dimethyl(methylthio)sulfonium tetrafluoroborate
- Phth
phthaloyl
- MBn
p-methoxybenzyl
- ClBn
p-chlorobenzyl 相似文献
6.
Guiyun Xu Takashi Suzuki Yasuhiro Maejima Tohru Mizoguchi Mamoru Tsuchiya Makoto Kiso Akira Hasegawa Yasuo Suzuki 《Glycoconjugate journal》1995,12(2):156-161
The sialidase of swine influenza A viruses of N1 and N2 subtypes, isolated from 1930 to 1992, was studied for substrate specificity with ganglio-series, lacto-series type II and GM3 gangliosides containing Neu5Ac2-3Gal, Neu5Gc2-3Gal and Neu5Ac2-6Gal linkages. All viral sialidases tested showed that the activity for hydrolysing substrates with Neu5Ac2-3Gal was higher than the activities with Neu5Gc2-3Gal and Neu5Ac2-6Gal linkages. When GM1b, GM3 and sialylparagloboside were used as substrates, the earliest strain (A/Wisconsin/15/30 H1N1, isolated in 1930) showed the activity ratio of Neu5Ac2-6Gal to Neu5Ac2-3Gal to be 0.13:0.2, and the ratio Neu5Gc2-3Gal/Neu5Ac2-3Gal to be 0.19:0.37, while those strains isolated from 1978 to 1992 exhibited ratios of 0.29:0.58 for Neu5Ac2-6Gal/Neu5Ac2-3Gal and 0.51:0.76 for Neu5Gc2-3Gal/Neu5Ac2-3Gal. The above results indicate that the substrate specificities of sialidases from swine influenza A viruses towards sialyl linkages and the molecular species of sialic acid are related to the year of isolation, i.e. strains isolated after 1978 exhibited higher activity towards Neu5Ac2-6Gal and Neu5Gc2-3Gal linkages when compared with strains isolated in an earlier year, 1930.Abbreviation Neu5Ac
5-N-acetylneuraminic acid
- Neu5Gc
5-N-glycolyneuraminic acid
- Gal
d-galactose
- Glc
d-glucose
- Cer
Ceramide
- II3(Neu5Ac)Lac
Neu5Ac2-3Gal1-4Glc
- GM3(Neu5Ac2-3Gal)
Neu5Ac2-3Gal1-4Glc1-Cer
- GM3(Neu5Gc2-3Gal)
Neu5Gc2-3Gal1-4Glc1-Cer
- GM1b(Neu5Ac2-3Gal)
Neu5Ac2-3Gal1-3GalNac1-4Gal1-4Glc1-Cer
- GMlb(Neu5Gc2-3Gal)
Neu5Gc2-3Gal1-3GalNAc1-4Gal1-4Glc1-Cer
- IV3(Neu5Ac)nLc4Cer
Neu5Ac2-3Gal1-3GlcNAc1-4Gal1-4Glc1-Cer
- IV3(Neu5Gc)nLc4Cer
Neu5Gc2-3Gal1-3GlcNAc1-4Gal1-4Glc1-Cer
- IV6(Neu5Ac)nLc4Cer
Neu5Ac2-6Gal1-3GlcNAc1-4Gal1-4Glc1-Cer
- TDC
taurodeoxycholate. 相似文献
7.
Cecilia PC Soh Alastair SR Donald James Feeney Walter TJ Morgan Winifred M Watkins 《Glycoconjugate journal》1989,6(3):319-332
The tetrasaccharides GalNAcß1-4[NeuAc2-3]Galß1-4Glc and GalNAcß1-4[NeuAc2-3]Galß1-4GlcNAc were synthesised by enzymic transfer of GalNAc from UDP-GalNAc to 3-sialyllactose (NeuAc2-3Galß1-4Glc) and 3-sialyl-N-acetyllactosamine (NeuAc2-3Galß1-4GlcNAc). The structures of the products were established by methylation and1H-500 MHz NMR spectroscopy. In Sda serological tests the product formed with 3-sialyl-N-acetyllactosamine was highly active whereas that formed with 3-sialyllactose had only weak activity. 相似文献
8.
N. Schilling 《Planta》1982,154(1):87-93
The de novo synthesis of maltose in spinach (Spinacia oleracea L.) was shown to be catalyzed by a maltose synthase, which converts two molecules of -d-glucose-1-phosphate (-G1P) (Km 1.5 mmol l-1) to maltose and 2 orthophosphate (Pi). This enzyme was purified 203-fold by fractionated ammonium sulfate precipitation and by column chromatography on Sepharose 6B. The addition of -G1P (15 mmol l-1) to the isolation buffer is required to stabilize the enzyme activity during the extraction and purification procedure. Molecular weight determination by gel filtration yielded a value of 95,000. -Gluconolactone, ATP and Pi are competitive inhibitors toward the substrate -G1P. The maltose synthase catalyzes an exchange of the phosphate group of -G1P with [32P] orthophosphate; this transfer reaction suggests that the synthesis of maltose occurs via a glucose-enzyme in a double displacement reaction. The physiological role of this enzyme as a starch initiator system is discussed.Abbreviations Fru
fructose
- Glc
glucose
- -G1P
-d-glucose-1-phosphate
- -G1P
-d-glucose-1-phosphate
- G6P
d-glucose-6-phosphate
This enzyme is tentatively called maltose synthase in this publication 相似文献
9.
Hirotaka Yamamoto Yoichi Kaneko David Vandermulen Donna Kersey Edward Mkrdichian Leonard Cerullo Jan Leestma Joseph R. Moskal 《Glycoconjugate journal》1995,12(6):848-856
The expression of CMP-NeuAc: Gal1,4GlcNAc 2,6 sialyltransferase (2,6-ST) [EC 2.4.99.1] and glycoproteins bearing 2,6-linked sialic acids were examined in primary human brain tumours and cell lines. 79% (19/24) of the meningiomas expressed 2,6-ST mRNA, 42% (10/24) of which showed very high expression. 2,6-ST mRNA expression was undetectable in normal brain tissue. In contrast, only 1/13 of the gliomas examined expressed detectable 2,6-ST mRNA. Metastases to the brain did not express measurable amounts of 2,6-ST mRNA. Less expression was found in malignant (i.e., anaplastic) compared to benign (i.e. meningothelial) meningiomas. Two-dimensional SDS-PAGE of glioma and meningioma proteins, followed bySambucus nigra lectin staining, revealed the presence of a glycoprotein bearing 2,6-linked sialic acids,M
r=53 kDa and a pI=7.0 (MEN-1) that appeared in all seven of the meningiomas examined, but was expressed at barely detectable levels, if at all, in seven out of the seven glioblastomas examined. Thus, decreased 2,6-ST expression may play a role in the aggressive nature of anaplastic meningiomas, but appears to be virtually absent in all tumours of glial origin. 相似文献
10.
Pascal Reboul Pascal George Delphine Miquel Pierre Louisot Pierre Broquet 《Glycoconjugate journal》1996,13(1):69-79
When treated with retinoic acidin vivo, C6 glioma cells show an enhancement of CMP-Neu5Ac:Gal 1–3 GalNAc-R -2,3 sialyltransferase activity. A 300kDa glycoprotein was detected by lectin affinoblotting in retinoic acid-treated C6 cells which stained weakly or not at all in control cells. Comparative studies with different lectins demonstrated that this glycoprotein contains 2,3 Neu5Ac Gal-GalNAc O-glycan moieties. Cultures in the presence of an inhibitor of O-glycan synthesis (N-acetylgalactosaminide -O-benzyl) demonstrated that enhancement of staining of the 300 kDa glycoprotein was not due to the increase of the 2,3 sialyltransferase but to thede novo synthesis of the polypeptide chain of this glycoprotein.Abbreviations RA
retinoic acid
- Neu5Ac
N-acetylneuraminic acid
- CMP-Neu5Ac
cytidine 5 monophosphosialate
- 2,3 ST
CMP-Neu5Ac:Gal 1–3 GalNAc-R -2,3 sialyltransferase
- GalNAc-O-benzyl
N-acetylgalactosaminide -O-benzyl
- Gal1-3GalNAc-O-benzyl
Galactosyl 1-3N-acetylgalactosaminide -O-benzyl
- TBS
Tris-HCl buffer 50mm pH 7.5 containing NaCl 0.15m and Tween 20 0.05%
- B1 buffer
TBS containing MgCl2 1mm, MnCl2 1mm and CaCl2 1mm 相似文献
11.
A rapid procedure is described for the separation of CMP-sialic acid:lactosylceramide sialyltransferase reaction components using Sep Pak C18 cartridges. The quantitative separation of the more polar nucleotide sugar, CMP-sialic acid, and its free acid from the less polar GM3-ganglioside is simple and rapid relative to previously described methods. Recovery of GM3 is optimized by the addition of phosphatidylcholine to the reaction mixture prior to the chromatographic step. Using rat liver Golgi membranes as a source of CMP-sialic acid: lactosylceramide sialyltransferase activity (GM3 synthase; ST-1), the transfer of [14C] sialic acid from CMP-[14C] sialic acid to lactosylceramide can be quantified by this assay. The procedure is reliable and may be applicable to the isolation of ganglioside products in otherin vitro glycosyltransferase assays.Abbreviations GM3
GM3-ganglioside
- II3NeuAc-LacCer
NeuAc2-3Gal1-4Glc1-1Cer
- GD1a
GD1a-ganglioside, IV3NeuAc, II3NeuAc-GgOse4Cer, NeuAc2-3Gal1-3GalNac1-4(NeuAc2-3)Gal1-4Glc1-1Cer
- GD3
GD3-ganglioside, II3(NeuAc)2LacCer, NeuAc2-8NeuAc2-3Gal1-4Glc1-1Cer
- GgOse4Cer
asialo-GM1 Gal1-3GalNAc1-4Gal1-4Glc1-1Cer
- FucGMI
fucosyl-GMI-ganglioside, Fuc1-2Gal1-3GalNAc1-4Gal1-4 Glc1-1Cer
- ST-1
GM3 synthase, CMP-sialic acid:lactosylceramide sialyltransferase
- LacCer
lactosylceramide, Gal1-4Glc1-1Cer
- CMP-NeuAc
cytidine 5-monophospho-N-acetylneuraminic acid
- PC
phosphatidylcholine
- PMSF
phenylmethylsulfonyl fluoride 相似文献
12.
The purified isopropylmalate synthase of Alcaligenes eutrophus H 16 reacted with the following -keto acids and acyl-coenzyme A derivatives (in the sequence of decreasing affinities): -ketoisovalerate, -keto-n-valerate, -ketobutyrate and pyruvate; acetyl-CoA, propionyl-CoA, butyryl-CoA. malonyl-CoA, valeryl-CoA, and crotonyl-CoA. -Ketoisocaproate, however, is a strong inhibitor of the enzyme. All reactions catalyzed by isopropylmalate synthase were inhibited to the same extent by the endproduct l-leucine. the substrate saturation curves of -ketoisovalerate or other -keto acids and of acetyl-coenzyme A or other acyl-CoA derivatives had intermediary plateau regions; the Hill coefficient alternated between n
H
-values higher and lower than 1.0, indicating changes from positive to negative and from negative to positive cooperativity for the substrates. The products, isopropylmalate and free coenzyme A, showed competitive inhibition patterns against both substrates (-ketoisovalerate and acetyl-CoA). Free coenzyme A (1 M) inactivated the enzyme irreversibly. The 3-phosphate of coenzyme A and the free carboxyl group of -ketoisovalerate were involved in optimal binding of these substrates, but 3-dephospho-acetyl-coenzyme A and the methylester of -ketoisovalerate were also converted by this enzyme. A CH3–CH2-grouping of the -keto acids seemed to be necessary for binding this substrate.Abbreviations Used CoA
Coenzyme A
- Tris
Tris(hydroxymethyl)aminomethane hydrochloride
- DTNB
5,5-dithiobis-(2-nitrobenzoic acid)
- IPM
-Isopropylmalate
- KIV
-Ketoisovalerate
Prepared from doctoral thesis of the University of Göttingen 1973 相似文献
13.
Summary A mutant strain of Rhodococcus equi accumulates three metabolites from the androst-4-ene-3,17-dione or from its degradation intermediate, 3a-H-4(3'-propionic acid)-7a-methylhexahydro-1,5-indanedione (MEPHIP). These three metabolites are: 3a-H-4a(3'-propionic acid)-5-hydroxy-7a-methylhexahydro-1-indanone--lactone (HIL); 3a-H-4(3'-trans acrylic acid)-5-hydroxy-7a-methylhexahydro-1-indanone (2'-5-hydroxy-MEPHIP); and 3a-H-4(3'-hydroxy-3'-propionic acid)-5-hydroxy-7a-methylhexahydro-1-indanone (3'-hydroxy-HIL). The behaviour of this mutant allows us to propose a pathway for degradation of the intermediates, methylperhydroindanone propionates. However, during this degradation, the side-chain propionate was eliminated by a-oxidation mechanism.
Offprint requests to: A. Miclo 相似文献
14.
Cornelis H. Hokke Jos G. M. van der Ven Johannis P. Kamerling Johannes F. G. Vliegenthart 《Glycoconjugate journal》1993,10(1):82-90
Incubation of synthetic Man\1-4GlcNAc-OMe, GalNAc1-4GlcNAc-OMe, Glc1-4GlcNAc-OMe, and GlcNAc1-4GlcNac-OMe with CMP-Neu5Ac and rat liver Gal1-4GlcNAc (2-6)-sialyltransferase resulted in the formation of Neu5Ac2-6Man1-4GlcNAc-OMe, Neu5Ac2-6GalNAc1-4GlcNAc-OMe, Neu5Ac2-6Glc1-4GlcNAc-OMe and Neu5Ac2-6GlcNAc1-4GlcNAc-OMe, respectively. Under conditions which led to quantitative conversion of Gal1-4GlcNAc-OEt into Neu5Ac2-6Gal1-4GlcNAc-OEt, the aforementioned products were obtained in yields of 4%, 48%, 16% and 8%, respectively. HPLC on Partisil 10 SAX was used to isolate the various sialyltrisaccharides, and identification was carried out using 1- and 2-dimensional 500-MHz1H-NMR spectroscopy.Abbreviations 2D
2-dimensional
- CMP
cytidine 5-monophosphate
- CMP-Neu5Ac
cytidine 5-monophospho--N-acetylneuraminic acid
- COSY
correlation spectroscopy
- DQF
double quantum filtered
- HOHAHA
homonuclear Hartmann-Hahn
- MLEV
composite pulse devised by M. Levitt
- Neu5Ac
N-acetylneuraminic acid
- Neu5Ac2en
2-deoxy-2,3-didehydro-N-acetylneuraminic acid 相似文献
15.
Summary Complex carbohydrates in the human cervix were studied histochemically using lectins, conjugated to horseradish peroxidase and correlated procedures. Stratified squamous epithelium of the exocervix and columnar epithelium of the endocervix in some, but not all specimens showed staining for terminal -N-acetyl-d-galactosamine, -d-galactose, -d-galactose and -l-fucose. the staining for -N-acetylgalactosamine and -galactose, the terminal sugars in blood group A and B antigens respectively, corresponded to a large extent with ABO blood type. One exception was the lack of staining for terminal -N-acetylgalactosamine in endocervical secretions in three of nine blood type A patients. A second exception was the staining for terminal -galactose in endocervical secretions in about half of blood type O and A specimens. The type and amount of glycoprotein formed by endocervical columnar cells differed according to location in superficial compared with deep portions of the glands and according to location at the junction with exocervix compared with the more internal regions. Staining of endothelial cells for blood group A and B antigens was confined to subjects of blood type A and B respectively, although three of nine type A specimens showed no lectin reactivity for group, A antigen. Endothelial cells evidenced affinity forUlex europeus I agglutinin demonstrative of fucose in all specimens. Mast cells disclosed lectin affinity consistent with the presence of terminal or internal mannose orN-acetylglucosamine residues. Two blood type O specimens were examined with conjugated lectins at the ultrastructural level. Secretory granules stained for content of terminal -galactose, -galactose and fucose. These results support and concur with biochemical studies of complex carbohydrates in human cervical tissues. They reveal, in addition, the location of the blood group antigens in the human exocervix and endocervix and the marked heterogeneity among endocervical columnar cells in glycoprotein production. 相似文献
16.
Summary Partitioning behaviour and adsorption isotherms of lactase/CM-Sephadex system at equilibrium were investigated together with the adsorption kinetics in this study. Maximum adsorption was obtained at the pH values between 5.5–6.0. Adsorption isotherm was a close fit to the Langmuir model.Nomenclature a
specific mass transfer area
- Dm
molecular diffusion coefficient (m2/sec)
- e1, e2
charge of the protein and the gel
- k
apparent mass transfer coefficient (s-1)
- ka
global mass transfer coefficient
- f
partition coefficient
- Kp
dissociation constant for adsorbent-adsorbate complex, (mg/mL solvent)
- p
equilibrium concentration of free enzyme, (mg free enzyme/mL solution)
- q
equilibrium concentration of adsorbed enzyme, (mg ads./mL gel)
- qm
maximum adsorption capacity, (mg ads./ml gel)
- Re
particle Reynolds number
- Sh
Sherwood number
- Vg/V
gel volume (mL)/bulk solvent volume (mL)
- Z
dimensionless extent of adsorption
-
Kp/Po , model parameter
-
(/) +1 , model parameter
-
Vg qm / V Po , model parameter 相似文献
17.
Jean-Michel Wieruszeski Jean-Claude Michalski Jean Montreuil Gérard Strecker 《Glycoconjugate journal》1990,7(1):13-26
The major pentasaccharides Fuc(1-2)[GalNAc(1-3)]Gal(1-4)[Fuc(1-3)]Glc and Fuc(1-2) [Gal(1-3)]Gal(1-4)[Fuc(1-3)]Glc, which are normally present in the urine of bloodgroup A Leb and B Leb healthy subjects, were each found to be contaminated by a minor component when analysed by1H-NMR. The determination of these structures, Fuc(1-2) [GalNAc(1-3)]Gal(1-3)[Fuc(1-4)]Glc and Fuc(1-2) [Gal(1-3)]Gal(1-3)[Fuc(1-4)]Glc, was based on the results of methylation analysis and1H/13C-NMR spectroscopy.Abbreviations HPLC
high performance liquid chromatography
- GLC
gas liquid chromatography
- NMR
nuclear magnetic resonance
- COSY
correlation spectroscopy
- Gal
d-galactopyranose
- GalNAc
2-acetamido-2-deoxy-d-galactopyranose
- Glc
d-glucopyranose
- Fuc
l-fucopyranose
- LNDFH I
lacto-N-difucohexaose I (Leb determinant 相似文献
18.
We examined the effects of changing KCl concentration on the secondary structures of -actinins using circular dichroism (CD), 1,1-bis(4-anilino) naphthalene-5,5-disulfonic acid (bisANS) fluorescence and proteolysis experiments. Under near-physiological conditions, divalent cations also were added and changes in conformation were investigated. In 25 mm KH2PO4, pH 7.5, increasing KCl from 0 to 120 mm led to decreases in -helix conformation for brain, platelet and heart -actinins (40.5-30.2%, 65.5-37.8% and 37.5-27.8%, respectively). In buffered 120 mm KCl, 0.65 mm calcium produced small changes in the CD spectra of both brain and platelet -actinin, but had no effect on heart -actinin. bisANS fluorescence of all three -actinins also showed significant changes in conformation with increasing KCl. However, in buffered 120 mm KCl increasing concentrations of Ca2+ or Mg2+ did not have significant effects on the bisANS fluorescence of any -actinin. Digestion of brain, platelet and heart -actinins with -chymotrypsin showed an increase of proteolytic susceptibility in 120 mm KCl. These experiments also showed that increasing the concentration of Ca2+ or Mg2+ led to greater changes in digestion fragment patterns in the absence of KCl than in the presence of 120 mm KCl. The results suggest that -actinins exist in different conformations depending on the ionic strength of the medium, which could explain the differences in calcium and F-actin binding results obtained from different -actinins. 相似文献
19.
Folkert Reck Ernst Meinjohanns Matthias Springer Roland Wilkens Johannes A. L. M. Van Dorst Hans Paulsen Gabriele Möller Inka Brockhausen Harry Schachter 《Glycoconjugate journal》1994,11(3):210-216
UDP-GlcNAc: Man1-6R (1-2)-N-acetylglucosaminyltransferase II (GlcNAc-T II; EC 2.4.1.143) is a key enzyme in the synthesis of complexN-glycans. We have tested a series of synthetic analogues of the substrate Man1-6(GlcNAc1-2Man1-3)Man-O-octyl as substrates and inhibitors for rat liver GlcNAc-T II. The enzyme attachesN-acetylglucosamine in 1-2 linkage to the 2-OH of the Man1-6 residue. The 2-deoxy analogue is a competitive inhibitor (K
i=0.13mm). The 2-O-methyl compound does not bind to the enzyme presumably due to steric hindrance. The 3-, 4- and 6-OH groups are not essential for binding or catalysis since the 3-, 4- and 6-deoxy and -O-methyl derivatives are all good substrates. Increasing the size of the substituent at the 3-position to pentyl and substituted pentyl groups causes competitive inhibition (K
i=1.0–2.5mm). We have taken advantage of this effect to synthesize two potentially irreversible GlcNAc-T II inhibitors containing a photolabile 3-O-(4,4-azo)pentyl group and a 3-O-(5-iodoacetamido)pentyl group respectively. The data indicate that none of the hydroxyls of the Man1-6 residue are essential for binding although the 2- and 3-OH face the catalytic site of the enzyme. The 4-OH group of the Man-O-octyl residue is not essential for binding or catalysis since the 4-deoxy derivative is a good substrate; the 4-O-methyl derivative does not bind. This contrasts with GlcNAc-T I which cannot bind to the 4-deoxy-Man- substrate analogue. The data are compatible with our previous observations that a bisectingN-acetylglucosamine at the 4-OH position prevents both GlcNAc-T I and GlcNAc-T II catalysis. However, in the case of GlcNAc-T II, the bisectingN-acetylglucosamine prevents binding due to steric hindrance rather than to removal of an essential OH group. The 3-OH of the Man1-3 is an essential group for GlcNAc-T II since the 3-deoxy derivative does not bind to the enzyme. The trisaccharide GlcNAc1-2Man1-3Man-O-octyl is a good inhibitor (K
i=0.9mm). The above data together with previous studies indicate that binding of the GlcNAc1-2Man1-3Man- arm of the branched substrate to the enzyme is essential for catalysis.
Abbreviations: GlcNAc-T I, UDP-GlcNAc:Man1-3R (1-2)-N-acetylglucosaminyltransferase I (EC 2.4.1.101); GlcNAc-T II, UDP-GlcNAc:Man1-6R (1-2)-N-acetylglucosaminyltransferase II (EC 2.4.1.143); MES, 2-(N-morpholino)ethane sulfonic acid monohydrate. 相似文献
20.
Hyaluronidase from the venom of the honeybee (Apis mellifera) has been purified by gelpermeation and cation exchange chromatography. Its asparagine-linked carbohydrate chains were released from tryptic glycopeptides with N-glycosidase A and reductively aminated with 2-aminopyridine. Separation of the fluorescent derivatives by size-fractionation and reversed-phase HPLC afforded eighteen fractions which were analysed by two-dimensional HPLC mapping combined with exoglycosidase digestions. The bulk of the N-linked glycans of hyaluronidase consisted of small oligosaccharides (Man1–3GlcNAc2), most of which were either 1,3-monofucosylated or 1,3-(1,6-)difucosylated at the innermost GlcNAc residue. High-mannose type structures constituted the minor fractions, together making up about 5% of the oligosaccharide pool from hyaluronidase. Four fractions, making up 8% of the N-linked glycans, contained the terminal trisaccharide GalNAc1-4[Fuc1-3]GlcNAc1- in 1,2-linkage to the core 1,3-mannosyl residue. No evidence for the presence of O-glycans or sialic acids could be found.Abbreviations GalNAc
N-acetylgalactosamine
- GlcNAc
N-acetylglucosamine
- PA
pyridylamino
- PLA
phospholipase A2
- 2D-HPLC
two-dimensional HPLC 相似文献