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91.
Stephen M. Henry Per-?ke Jovall Sohbat Ghardashkhani Mikael L. Gustavsson Bo E. Samuelsson 《Glycoconjugate journal》1995,12(3):309-317
Total non-acid glycosphingolipids were isolated from the plasma of a healthy red blood cell group O Le(a-b-) salivary ABH secretor individual. Glycolipids were fractionated by HPLC and combined into eight fractions based on chromatographic and immunoreactive properties. These glycolipid fractions were analysed by thin-layer chromatography and tested for Lewis activity with antibodies reactive to the type 1 precursor (Lec), H type 1 (Led), Lea and Leb epitopes. Fractions were structurally characterized by mass spectrometry (EI-MS and LSIMS) and proton NMR spectroscopy. Expected blood group glycolipids, such as H type 1, (Fuc1-2Gal1-3GlcNAc1-3Gal1-4Glc1-1Cer) were immunochemically and structurally identified. Inconsistent with the red cell phenotype and for the first time, small quantities of Leb blood group glycolipids (Fuc1-2Gal1-3(Fuc1-4)GlcNAc1-3Gal1-4Glc1-1Cer) were immunochemically and structurally identified in the plasma of a Lewis-negative individual. These findings confirm recent immunological evidence suggesting the production of small amounts of Lewis antigens by Lewis negative individuals.
Abbreviations: HPLC, high performance liquid chromatography; TLC, (high performance) thin layer chromatography; EI-MS, electron impact ionisation mass spectrometry; LSIMS, liquid secondary ion mass spectrometry; NMR, nuclear magnetic resonance spectroscopy. The sugar types are abbreviated to Hex for hexose, HexNAc forN-acetylhexosamine and dHex for deoxyhexose (fucose). The ceramide types are abbreviated to d for dihydroxy and t for trihydroxy base, n for non-hydroxy and h for hydroxy fatty acids; LCB, long chain base. 相似文献
92.
Edouard Nice Bruno Catimel Martin Lackmann Steven Stacker Andrew Runting Andrew Wilks Nicos Nicola Antony Burgess 《Letters in Peptide Science》1997,4(2):107-120
The isolation of related genes with evolutionary conserved motifs by the application ofpolymerase chain reaction-based molecular biology techniques, or from database searchingstrategies, has facilitated the identification of new members of protein families. Many of theseprotein molecules will be involved in protein–protein interactions (e.g. growth factors,receptors, adhesion molecules), since such interactions are intrinsic to virtually every cellularprocess. However, the precise biological function and specific binding partners of these novelproteins are frequently unknown, hence they are known as orphan molecules.Complementary technologies are required for the identification of the specific ligands orreceptors for these and other orphan proteins (e.g., antibodies raised against crude biologicalextracts or whole cells). We describe herein several alternative strategies for the identification,purification and characterisation of orphan peptide and protein molecules, specifically thesynergistic use of micropreparative HPLC and biosensor techniques. 相似文献
93.
Qinghong Xu Elizabeth A. Ottinger Nuria A. Solé George Barany 《Letters in Peptide Science》1997,3(6):333-342
Summary In the course of solid-phase synthesis of phosphopeptides by a post-assembly global phosphorylation strategy, the corresponding H-phosphonate peptides form as byproducts. We describe model studies to investigate this side reaction as a function of reaction conditions, and use this information to develop conditions that minimize the problem, i.e., use of dibenzyl N,N-di-isopropyl phosphoramidite for phosphitylation, followed immediately by oxidation with anhydrous tert-butyl hydroperoxide in dry tetrahydrofuran under argon, and final acidolytic cleavage.This work was taken in part from the Ph.D. Theses of E.A. Ottinger (1994) and Q. Xu (1996), University of Minnesota, Minneapolis, MN, U.S.A. Preliminary presentations of portions of this work were made at the Twenty-Second European Peptide Symposium, Interlaken, Switzerland, September 13–19, 1992, see Ref. 1, at the 14th American Peptide Symposium, Columbus, OH, U.S.A., June 18–23, 1995, and at the Fourth International Symposium on Solid Phase Synthesis & Combinatorial Chemical Libraries, Edinburgh, Scotland, U.K., September 12–16, 1995, see Ref. 2. The title side reaction was first discussed for tyrosine (see Refs 1 and 3), but all of the mechanism studies discussed herein are for serine and threonine.Amino acid symbols denote the l-configuration, and abbreviations for amino acids and peptides follow rules of the IUPAC-IUB Commission of Biochemical Nomenclature [J. Biol. Chem., 247 (1972) 977]. 相似文献
94.
Mikkel Nissum Jens-Jakob Karlsson Jens Ulstrup Palle Waage Jensen G. Smulevich 《Journal of biological inorganic chemistry》1997,2(3):302-307
Di-heme Pseudomonas stutzeri cytochrome c
4 has been characterized by electronic absorption and resonance Raman spectroscopies in the ferric and ferrous forms at pH 7.5
and at room temperature. The data indicate that the two hemes are inequivalent. It is proposed that the N-terminal contains
a more relaxed heme as a consequence of the relative orientation of the methionine and histidine ligands with respect to the
N-Fe-N directions of the heme plane. This causes a weakening of the Fe-S bond with concomitant partial dissociation of the
methionine and the formation of an Fe-aquo bond. Heme group relaxation is further accompanied by less distortion of the heme
group than that associated with cytochrome c, expansion of the "core" and a negative shift of the redox potential.
Received: 17 December 1996 / Accepted: 6 March 1997 相似文献
95.
Complete 1H, 13C, and 15N NMR resonance assignments and secondary structure of human glutaredoxin in the fully reduced form. 下载免费PDF全文
C. Sun A. Holmgren J. H. Bushweller 《Protein science : a publication of the Protein Society》1997,6(2):383-390
Human glutaredoxin is a member of the glutaredoxin family, which is characterized by a glutathione binding site and a redox-active dithiol/disulfide in the active site. Unlike Escherichia coli glutaredoxin-1, this protein has additional cysteine residues that have been suggested to play a regulatory role in its activity. Human glutaredoxin (106 amino acid residues, M(r) = 12,000) has been purified from a pET expression vector with both uniform 15N labeling and 13C/15N double labeling. The combination of three-dimensional 15N-edited TOCSY, 15N-edited NOESY, HNCA, HN(CO)CA, and gradient sensitivity-enhanced HNCACB and HNCO spectra were used to obtain sequential assignments for residues 2-106 of the protein. The gradient-enhanced version of the HCCH-TOCSY pulse sequence and HCCH-COSY were used to obtain side chain 1H and 13C assignments. The secondary structural elements in the reduced protein were identified based on NOE information, amide proton exchange data, and chemical shift index data. Human glutaredoxin contains five helices extending approximately from residues 4-10, 24-36, 53-64, 83-92, and 94-104. The secondary structure also shows four beta-strands comprised of residues 15-19, 43-48, 71-75, 78-80, which form a beta-sheet almost identical to that found in E. coli glutaredoxin-1. Complete 1H, 13C, and 15N assignments and the secondary structure of fully reduced human glutaredoxin are presented. Comparison to the structures of other glutaredoxins is presented and differences in the secondary structure elements are discussed. 相似文献
96.
The Lipid Peroxidation Product, 4-Hydroxy-2-trans-Nonenal, Alters the Conformation of Cortical Synaptosomal Membrane Proteins 总被引:6,自引:1,他引:5
‡Ram Subramaniam ‡Fred Roediger Brad Jordan †‡§Mark P. Mattson §Jeffrey N. Keller Georg Waeg ‡§ D. Allan Butterfield 《Journal of neurochemistry》1997,69(3):1161-1169
Abstract: Alzheimer's disease (AD) is widely held to be a disorder associated with oxidative stress due, in part, to the membrane action of amyloid β-peptide (Aβ). Aβ-associated free radicals cause lipid peroxidation, a major product of which is 4-hydroxy-2- trans -nonenal (HNE). We determined whether HNE would alter the conformation of synaptosomal membrane proteins, which might be related to the known neurotoxicity of Aβ and HNE. Electron paramagnetic resonance spectroscopy, using a protein-specific spin label, MAL-6(2,2,6,6-tetramethyl-4-maleimidopiperidin-1-oxyl), was used to probe conformational changes in gerbil cortical synaptosomal membrane proteins, and a lipid-specific stearic acid label, 5-nitroxide stearate, was used to probe for HNE-induced alterations in the fluidity of the bilayer domain of these membranes. Synaptosomal membranes, incubated with low concentrations of HNE, exhibited changes in protein conformation and bilayer order and motion (fluidity). The changes in protein conformation were found to be concentration- and time-dependent. Significant protein conformational changes were observed at physiologically relevant concentrations of 1–10 µ M HNE, reminiscent of similar changes in synaptosomal membrane proteins from senile plaque- and Aβ-rich AD hippocampal and inferior parietal brain regions. HNE-induced modifications in the physical state of gerbil synaptosomal membrane proteins were prevented completely by using excess glutathione ethyl ester, known to protect neurons from HNE-caused neurotoxicity. Membrane fluidity was found to increase at higher concentrations of HNE (50 µ M ). The results obtained are discussed with relevance to the hypothesis of Aβ-induced free radical-mediated lipid peroxidation, leading to subsequent HNE-induced alterations in the structure and function of key membrane proteins with consequent neurotoxicity in AD brain. 相似文献
97.
Hideyuki Takahashi Mitsuharu Inaki Koichi Fujimoto Shigeru Katsuta Izumi Anno Mamoru Nütsu Yuji Itai 《European journal of applied physiology and occupational physiology》1995,71(5):396-404
We examined the effect of differences in exercise intensity on the time constant (t
c) of phosphocreatine (PCr) resynthesis after exercise and the relationships betweent
c and maximal oxygen uptake (VO2max) in endurance-trained runners (n = 5) and untrained controls (n = 7) (average VO2max = 66.2 and 52.0 ml · min–1 · kg–1, respectively). To measure the metabolism of the quadriceps muscle using phosphorus nuclear magnetic resonance spectroscopy, we developed a device which allowed knee extension exercise inside a magnet. All the subjects performed four types of exercise: light, moderate, severe and exhausting. The end-exercise PCr: [PCr + inorganic phosphate (Pi)] ratio decreased significantly with the increase in the exercise intensity (P < 0.01). Although there was little difference in the end-exercise pH, adenosine diphosphate concentration ([ADP]) and the lowest intracellular pH during recovery between light and moderate exercise, significant changes were found at the two higher intensities (P < 0.01). These changes for runners were smaller than those for the controls (P < 0.05). The
c remained constant after light and moderate exercise and then lengthened in proportion to the increase in intensity (P < 0.05). The runners had a lowert
c at the same PCr and pH than the controls, particularly at the higher intensity (P < 0.05). There was a significant correlation betweent
c and [ADP] in light exercise and betweent
c and both end-exercise PCr and pH in severe and exhausting exercise (P < 0.05). The threshold of changes in pH andt
c was a PCr: (PCr + Pi) ratio of 0.5. There was a significant negative correlation between the VO2max andt
c after all levels of exercise (P<0.05).However, in the controls a significant correlation was found in only light and moderate exercise (P < 0.05). These findings suggest the validity of the use oft
c at an end-exercise PCr:(PCr + Pi) ratio of more than 0.5 as a stable index of muscle oxidative capacity and the correlation between local and general aerobic capacity. Moreover, endurance-trained runners are characterized by the faster PCr resynthesis at the same PCr and intracellular pH. 相似文献
98.
N. C. Hall R. J. Dempsey J. M. Carney D. L. Donaldson D. A. Butterfield 《Neurochemical research》1995,20(10):1161-1169
We have previously reported that ischemia reperfusion injury results from free radical generation following transient global ischemia, and that this radical induced damage is evident in the synaptosomal membrane of the gerbil. [Hall et al, (1995) Neuroscience 64: 81–89] In the present study we have extended these observations to transient focal ischemia in the cat. We prepared synaptosomal membranes from frontal, parietal-temporal, and occipital regions of the cat cerebral cortex with reperfusion times of 1 and 3 hours following 1 hour right middle cerebral artery occlusion. The membranes were selectively labeled with protein and lipid specific paramagnetic spin labels and analyzed using electron paramagnetic resonance spectrometry. There were significant motional changes of both the protein and lipid specific spin labels in the parietal-temporal and occipital regions with 1 hour reperfusion; but, both parameters returned to control values by 3 hours reperfusion. No significant changes were observed in the normally perfused frontal pole at either reperfusion time. These results support the argument that free radicals play a critical role in cell damage at early reperfusion times following ischemia. 相似文献
99.
S. S. Wijmenga H. A. Heus H. A. E. Leeuw H. Hoppe M. van der Graaf C. W. Hilbers 《Journal of biomolecular NMR》1995,5(1):82-86
Summary A new 1H−13C−31P triple resonance experiment is described which allows unambigous sequential backbone assignment in 13C-labeled oligonucleotides via through-bond coherence transfer from 31P via 13C to 1H. The approach employs INEPT to transfer coherence from 31P to 13C and homonuclear TOCSY to transfer the 13C coherence through the ribose ring, followed by 13C to 1H J-cross-polarisation. The efficiencies of the various possible transfer pathways are discussed. The most efficient route
involves transfer of 31Pi coherence via C4′i and C4′i-1, because of the relatively large J′PC4 couplings involved. Via the homonuclear and heteronuclear mixing periods, the C4′i and C4′i-1 coherences are subsequently transferred to, amongst others, H1′i and H1′i-1, respectively, leading to a 2D 1H−31P spectrum which allows a sequential assignment in the 31P−1H1′ region of the spectrum, i.e. in the region where the proton resonances overlap least. The experiment is demonstrated on
a 13C-labeled RNA hairpin with the sequence 5′(GGGC-CAAA-GCCU)3′. 相似文献
100.