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1.
The uptake and trapping constants for labeled tryptophan (Trp) via the serotonin (5-hydroxytryptamine; 5-HT) metabolic pathway and for the incorporation of Trp into proteins, and -[14C]methyl-L-tryptophan (-MTrp) were measured. Measurements were done in rats treated with either saline or probenecid (200 mg/kg). In addition, the blood-brain barrier (BBB) permeability surface area products for Trp (PST) and -MTrp (PS) were measured in normal rats. The results suggest that, in both groups of rats, there is a highly significant correlation (p < 0.05; Pearson Product Moment Correlation (PPMC) between the brain uptake and trapping constants for -MTrp and those of Trp via the 5-HT metabolic pathway, but there is no significant correlation (p > 0.05; PPMC) between either of these constants and the PS products of either compound. There is also no significant correlation (p > 0.05; PPMC) between the constant for the Trp incorporation into proteins with any of the other parameters. For all parameters, except Trp incorporation into proteins (-MTrp is not incorporated into proteins), there was a highly significant correlation (p < 0.001) between the quantities measured for Trp and -MTrp. The data presented here strongly suggests that the brain uptake and trapping of -MTrp relates to brain 5-HT synthesis, and does not relate to the BBB transport or protein incorporation of Trp. On the basis of these results, as well as those previously reported, we concluded that trapping (unidirectional uptake) of -MTrp can be converted to the 5-HT synthesis rates in the brain. From this also follows that labeled -MTrp is a good tracer for in vivo evaluation of the brain 5-HT synthesis. 相似文献
2.
The α/β Interfaces of α1β1, α3β3, and F1: Domain Motions and Elastic Energy Stored during γ Rotation
ATP synthase (FoF1) consists of F1 (ATP-driven motor) and Fo (H+-driven motor). F1 is a complex of 33 subunits, and is the rotating cam in 33. Thermophilic F1 (TF1) is exceptional in that it can be crystallized as a monomer and an 33 oligomer, and it is sufficiently stable to allow refolding and reassembly of hybrid complexes containing 1, 2, and 3 modified or . The nucleotide-dependent open–close conversion of conformation is an inherent property of an isolated and energy and signals are transferred through / interfaces. The catalytic and noncatalytic interfaces of both mitochondrial F1 (MF1) and TF1 were analyzed by an atom search within the limits of 0.40 nm across the interfaces. Seven (plus thermophilic loop in TF1) contact areas are located at both the catalytic and noncatalytic interfaces on the open form. The number of contact areas on closed increased to 11 and 9, respectively, in the catalytic and noncatalytic interfaces. The interfaces in the barrel domain are immobile. The torsional elastic strain applied through the mobile areas is concentrated in hinge residues and the P-loop in . The notion of elastic energy in FoF1 has been revised. X-ray crystallography of F1 is a static snap shot of one state and the elastic hypotheses are still inconsistent with the structure, dyamics, and kinetics of FoF1. The domain motion and elastic energy in FoF1 will be elucidated by time-resolved crystallography. 相似文献
3.
The C chemical shift tensors of proteins contain information on the backbone conformation. We have determined the magnitude and orientation of the C chemical shift tensors of two peptides with -helical torsion angles: the Ala residue in G*AL (=–65.7°, =–40°), and the Val residue in GG*V (=–81.5°, =–50.7°). The magnitude of the tensors was determined from quasi-static powder patterns recoupled under magic-angle spinning, while the orientation of the tensors was extracted from C–H and C–N dipolar modulated powder patterns. The helical Ala C chemical shift tensor has a span of 36 ppm and an asymmetry parameter of 0.89. Its 11 axis is 116° ± 5° from the C–H bond while the 22 axis is 40° ± 5° from the C–N bond. The Val tensor has an anisotropic span of 25 ppm and an asymmetry parameter of 0.33, both much smaller than the values for -sheet Val found recently (Yao and Hong, 2002). The Val 33 axis is tilted by 115° ± 5° from the C–H bond and 98° ± 5° from the C–N bond. These represent the first completely experimentally determined C chemical shift tensors of helical peptides. Using an icosahedral representation, we compared the experimental chemical shift tensors with quantum chemical calculations and found overall good agreement. These solid-state chemical shift tensors confirm the observation from cross-correlated relaxation experiments that the projection of the C chemical shift tensor onto the C–H bond is much smaller in -helices than in -sheets. 相似文献
4.
High frequency of triplicated α-globin loci and absence or low frequency of α thalassemia in Polynesian Samoans 总被引:3,自引:0,他引:3
Summary Most of the population in certain areas of Melanesia have one -globin gene deletion ( thal2). It is thought that the high frequencies of thal2 in this population is due to a selective advantage given by malaria infection to carriers of thal2. We are interested in neighboring Polynesia which, although adjacent to Melanesia, has always been free of malaria due to the absence of the vector anopheles. We studied 60 Polynesian Samoans and 150 Malaysians by restriction endonuclease gene mapping using Eco RI, Bam HI, and Bgl II and hybridization to 32P-labeled -globin gene probe. Seven among the 60 (11.7%) Samoans had triplicated -globin loci type 1, while none had thal2. On digestion with Bgl II the third -globin gene was found in an additional 3.7kb fragment in all seven Samoans with triplicated -globin loci, while digestion with Bam HI produced an abnormal elongated 18.2 kb fragment carrying -globin genes in addition to the normal 14.5 kb fragment. None of the Polynesian Samoans had thal2 or thal1. Only two of the Malaysians had triplicated -globin loci. 相似文献
5.
Giesen AW Bae LC Barrett CL Chyba JA Chaykovsky MM Cheng MC Murray JH Oliver EJ Sullivan SM Brown JM Homans SW 《Journal of biomolecular NMR》2002,22(1):21-26
The implementation of [13C,13C,15N,2H] labelled amino acids into proteins allows the acquisition of high resolution triple resonance experiments. We present for the first time resonance assignments facilitated by this new labelling strategy. The absence of 1JC,C couplings enables us to measure 1JC,C scalar and 1DC,C residual dipolar coupling constants using modified HNCA experiments which do not suffer from sensitivity losses characteristic for 13C constant time experiments. 相似文献
6.
Chimeric G proteins, obtained by exchanging their C-terminal portion for that of a G protein from an unrelated class, drive the receptor selectivity to that corresponding to the introduced G protein domain. The 2A-adrenoceptor (2AAR), which yielded an efficacious and weak [35S]GTPS binding response by respectively G o and G i3 protein, was investigated in CHO-K1 cells co-expressing chimeric G proteins for which the six last C-terminal amino acids between G o and G i3 proteins, and reciprocally, were permuted. Activation of the chimeric G o / i3 protein was highly efficient whereas the G i3 / o protein yielded a weak stimulation. These [35S]GTPS binding responses were not different from their parental wild-type G o and G i3 proteins. Similar results were obtained with an 2AAR carrying a facilitating Thr373Lys mutation in a putative G protein interaction domain. These data indicate that the six terminal G o protein amino acids do not constitute a major 2AAR interaction domain for G protein activation. 相似文献
7.
Mikala G Bodi I Klockner U Varadi M Varadi G Koch SE Schwartz A 《Molecular and cellular biochemistry》2003,250(1-2):81-89
The carboxyl terminal of the L-type calcium channel 1C subunit comprises approximately one third of the primary structure of the 1 subunit (> 700 amino acids residues). This region is sensitive to limited posttranslational processing. In heart and brain the 1C subunits are found to be truncated but the C-terminal domain remains functionally present. Based on our previous data we hypothesized that the distal C-terminus (approximately residues 1650–1950) harbors an important, predominantly inhibitory domain. We generated C-terminal-truncated 1C mutants, and after expressing them in combination with a 3 subunit in HEK-293 cells, electrophysiological experiments were carried out. In order to dissect the important inhibitory part of the C-terminus, trypsin was dialyzed into the cells. The data provide evidence that there are multiple residues within the inhibitory domain that are crucial to the inhibitory process as well as to the enhancement of expressed current by intracellular application of proteases. In addition, the expression of the chimeric mutant 1C1673-DRK1 demonstrated that the C-terminal is specific for the heart channel. 相似文献
8.
Philippe Pelupessy Elisabetta Chiarparin Ranajeet Ghose Geoffrey Bodenhausen 《Journal of biomolecular NMR》1999,13(4):375-380
The angle CH,NHN subtended by the internuclear vectors 13C-H and 15N-HN in doubly-labeled proteins can be determined by observing the effect of cross-correlation between the dipolar interactions on zero- and double-quantum coherences involving 13C and 15N. Two complementary 2D experiments with the appearance of 15N-HN correlation spectra yield signal intensities that depend on the rate of interconversion through cross-correlated relaxation of in-phase and doubly antiphase zero- and double-quantum coherences. The ratio of the signal intensities in the two experiments bears a simple relationship to the cross-correlation rate, and hence to the angle CH,NHN. Assuming planarity of the peptide bond, the dihedral angle (between C and C) can be determined from the knowledge of CH,NHN. The experiments are very time-effective and provide good sensitivity and excellent spectral resolution. 相似文献
9.
The Ca2+ channel 1B subunit is a pore-forming component capable of generating N-type Ca2+ channel activity. Although N-type Ca2+ channel plays a role in a variety of neuronal functions, 1B-deficient mice exhibit normal life span without apparent abnormalities of behavior, histology or plasma norepinephrine level, presumably owing to compensation by some other Ca2+ channel 1 or subunit. In this study, we studied the levels of 1A, 1C, 1D, 1E, 1, 2, 3 and 4 mRNAs in adrenal gland of 1B-deficient mice. The 1A mRNA in homozygous mice was expressed at higher level than in wild or heterozygous mice, but no difference in the expression levels of 1C, 1D, 1E, 1, 2, 3 and 4 was found among wild, heterozygous and homozygous mice. The protein level of 1A in homozygous mice was also expressed at higher level than in wild or heterozygous mice. To examine whether increased expression is induced by cis-regulatory element within 5-upstream region of 1A gene, we examined lacZ expression in 1B-deficient × 1A6.3-lacZ mice (carrying a 6.3-kb 5-upstream fragment of 1A gene fused to E. coli lacZ reporter gene), which express lacZ in medullar chromaffin cells, but not in cortex. The levels of lacZ expression in homozygous 1B-deficient × 1A6.3-lacZ mice were higher than in wild or heterozygous mice. Therefore, a possible explanation of the normal behavior and plasma norepinephrine level of 1B-deficient mice is that compensation by 1A subunit occurs and that 6.3-kb 5-upstream region of 1A gene contains enhancer cis-element(s) for compensation in adrenal medulla chromaffin cells. (Mol Cell Biochem 271: 91–99, 2005) 相似文献
10.
We studied the subunit composition of nicotinic cholinoreceptors (nChR) of neurons of the inferior mesenteric ganglion (IMG) of the guinea pig using antibodies against the 3, 4, 5, and 7 nChR subunits and a standard technique of intracellular recording. Application of the 3 subunit-specific antibodies resulted in a decrease in the EPSP amplitude by 29.7 ± 1.8%, on average, in 17 of 20 studied neurons. The 4 subunit-specific antibodies evoked no changes in the amplitude of EPSP recorded from IMG neurons (n = 10). Effects of the 5 subunit-specific antibodies were studied in 20 neurons, where the EPSP amplitude dropped after application of these antibodies by 40.0 ± 1.8%, on average. Superfusion of the neurons with a solution containing the 3 and 5 subunit-specific antibodies completely suppressed synaptic responses (n = 3). The 7 subunit-specific antibodies provided an increase in the EPSP amplitude in 13 of 15 studied IMG neurons (by 37 ± 6%, on average); this fact allows us to suppose that 7-containing nChR are present in the IMG neurons and are indirectly involved in the processes of synaptic transmission. Application of the antibodies evoked no significant shifts in the membrane potential in IMG neurons. Our findings demonstrate that synaptic transmission through the guinea pig IMG is realized mostly with the involvement of the 3- and 5-containing nChR; the 4-containing receptors are not engaged in this process. 相似文献
11.
Eric Mossel Fernando Formaggio Giovanni Valle Marco Crisma Claudio Toniolo Mitsunobu Doi Toshimasa Ishida Quirinus B. Broxterman Johan Kamphuis 《Letters in Peptide Science》1998,5(2-3):223-225
In order to obtain further information on the role played by phenyl ring position in the C-methylated -amino acid side chain on peptide preferred conformation, the crystal-state structural preferences of C-methyl, C-phenylglycine peptides have been determined by X-ray diffraction. This study shows that either the fully extended conformation or the -bend/310-helical structures are adopted by peptides characterized by this C-methylated, -branched, aromatic -amino acid. 相似文献
12.
13.
Sebastian Voigt Reinhart Gossrau Oliver Baum Klemens L?ster Werner Hofmann Werner Reutter 《The Histochemical journal》1995,27(2):123-132
Summary The 11-integrin is known to be a receptor for collagen and laminin mediating cell-matrix interactions. A monoclonal antibody, 33.4, which specifically inhibits the 1-integrin-mediated in vitro cell-collagen binding of rat hepatocytes and hepatoma-derived A-cells (Löster et al., 1994), was used to purify by immunoaffinity chromatography the 1-integrin subunit from rat liver in large quantities for inducing a polyclonal antiserum. In immunoblot analysis on membrane extracts of several rat organs this polyclonal antiserum recognized only a 190 kDa-band, suggesting that it is highly specific for the 1-integrin subunit. A sandwich-ELISA with monoclonal antibody 33.4 and the polyclonal antiserum against the 1-integrin subunit, respectively, enabled the quantitative expression pattern of the 1-integrin subunit to be studied in different rat organs. With the exceptions of brain (not detectable) and muscle (low concentration), the 1-integrin subunit was detectable in almost all organs of the digestive, respiratory and urogenital system as well as in lymphatic organs. The highest relative concentrations of 1-integrin subunit were found in uterus, lung and spleen, whereas in seminal vesicle, stomach, parotid gland, epididymis, kidney and liver only modest concentrations were evident. The organ distribution and localization of 1-integrin subunit were studied by immunohistochemistry with monoclonal and polyclonal antibodies. Immunoreactivity was present in the plasma membranes of all smooth muscle cells, vascular endothelial cells of many organs and fibrocyte-fibroblast sheaths in the heart and kidney. Since these cells are in close contact with collagen-containing basal membranes as well as reticular fibrils, strong evidence exists that in rat tissues the 1-integrin subunit is expressed at sites where collagen is present and might be involved in vivo in cell—ollagen binding.Dedicated to Professor Peter Sitte on the occasion of his 65th birthday 相似文献
14.
Since, linking of ovine luteinizing hormone (oLH) to ribosome inactivating protein gelonin (in oLH-gelonin conjugate) occur via the alpha-subunit, oLH, an attempt has been made to develop a universal hormonotoxin for selective targeting to specific cells in the gonads. Four different molar ratios of oLH and N-succinimidyl-3-(2-pyridyldithio) propionate (SPDP) were used to activate the epsilon amino (-NH2) groups of oLH. The oLH-SPDP derivatives recombine to native beta subunit of oLH (oLH) and the purified recombinants retained substantial receptor binding, steroidogenic activity and immunoreactivity to native oLH. The disulfide linked oLH-S-S-gelonin conjugates prepared by SPDP method were purified by gel filtration chromatography and analysed by reverse-phase high performance liquid chromatography (RP-HPLC). In order to obtain specificity and bioeffectivity, the oLH-S-S-gelonin conjugates were allowed to recombine to native oLH and the recombination mixture was further purified by gel-filtration chromatography. The RP-HPLC analysis of these recombinants indicated that oLH-S-S-gelonin did not recombine to oLH. The failure of recombination may be due to the reasons. (i) The site of -NH2 activation by SPDP may be different in the oLH than the native oLH. (ii) The activation site may be in close proximity to the annealing site which facilitates the recombination of -subunit but failured to reassociate to oLH-S-S-gelonin conjugate. (iii) The introduction of gelonin (30 kDa basic protein) might have induced some steric hinderence for oLH to recombine to the oLH site which might have been masked in oLH-S-S-gelonin conjugates. (Mol Cell Biochem120: 95–102, 1993)Abbreviations oLH
ovine Luteinizing Hormone
- oLH
alpha subunit of oLH
- oLH
beta subunit of oLH
- BSA
Bovine Serum Albumin
- DTT
Dithiothreitol
- RP-HPLC
Reverse Phase High Performance Liquid Chromatography
- TSH
Thyroid Stimulating Hormone
- FSH
Follicle Stimulating Hormone
- LH
Luteinizing Hormone
- eCG
equine Chorionic Gonadotropin
- DMEM
Dulbecco's Modified Eagles Medium
- HEPES
4-(2-Hydroxyethyl)-1 Piperazine Ethane Sulfonic acid
- PAP
Pokeweed Antiviral Protein
- RIA
Radioimmunoassay
- hCG
human Chorionic Gonadotropin
- TRH
Thyrotropin Releasing Hormone
- CRF
Corticotropin Releasing Factor
- hPL
human Placental Lactogen
- TFA
Trifluroacetic Acid
- oLH-SPDP
SPDP activated derivative of oLH 相似文献
15.
Li Guangyu Laabich Aicha Liu L. Olivia Xue Jin Cooper Nigel G.F. 《Molecular biology reports》2001,28(1):35-41
The aims of this study were to characterize specific mRNAs and the expression pattern for isoforms of calcium/calmodulin-dependent protein kinase II (CaMKII) in the human brain. We cloned and sequenced the CaMKII and subunit cDNAs, and used them to study the CaMKII expression in human brain. Four distinct isoforms of CAMKII were isolated. Two of them were characterized as CaMKII and subunits. The other two showed similar nucleotide sequences, but one had a 33-bp insertion relative to the subunit, and the other had a 75-bp deletion relative to the subunit. These alterations are located within the variable regions. These two isoforms were characterized as CaMKII B and e. Northern blot analysis showed that a 4.4-kb messenger RNA for the isoform and a 3.9-kb messenger RNA for the isoform were expressed in both human fetal and adult brain to different degrees. The results indicate that CaMKII expression is developmentally regulated. The CaMKII isoform expression was confirmed in human fetal and adult brain using RT-PCR with specific primers, which flanked the CaMKII variable regions. The CaMKII , B, , and e isoforms were characterized in both human fetal and adult brain. 相似文献
16.
Hitoshi Nakashima Asao Fujiyama Shunjiro Kagiyama Takashi Imamura 《Human genetics》1990,84(6):568-570
Summary Of the 645 Japanese subjects studied, we have identified 10 individuals heterozygous for a chromosome with the triplicated -globin loci. The frequency of the triple -loci was 0.008 in this population, while that of the single -locus, i.e., -thalassemia 2 gene, might be lower than 0.0008. Analysis of haplotypes using particular RsaI site polymorphism in the -globin gene complex strongly suggests that the triple loci may have had multiple origins in this population. 相似文献
17.
The nonproteinogenic amino acid, cyclopentenylglycine, is found in certain Flacourtiaceae. This compound may be synthesized by two C1-chain elongations of -ketoglutarate via -ketopimelate (C5+2C1) or by condensation of C4 and C3 units (C4+C3), a pathway not involving -ketopimelate. The following experimental design elucidated the biosynthetic pathway: Idesia polycarpa callus cultures were freshly established from leaf petioles; synthetic -[1,2-14C]ketopimelate was added to the medium and cultures were incubated for 3 weeks. After isolation and separation of free amino acids from the tissues, the radioactivity incorporated into cyclopentenylglycine was determined. The results establish -ketopimelate as a precursor for cyclopentenylglycine, thus providing evidence for the C5+2C1 biosynthetic path. 相似文献
18.
Jacques Raymond Brahim Mimouni Jean-Louis Azanza 《Plant Systematics and Evolution》1994,193(1-4):69-79
The seed storage globulins from sixHelianthus and four hybrids were studied using mono and bidimensional gel SDS electrophoresis (+ 2 mercaptoethanol). The polypeptide composition of each subunit was determined. Different pairs are specifically expressed according to the species studied. Three typical patterns were discriminated. All the studied species exhibit five subunits: two of them are expressed in all the species (11 and 22). The subunit corresponding to the 11 pair is present inH. petiolaris and in the three populations ofH. annuus studied. The 2b2 pair is common toH. annuus andH. argophyllus. H. petiolaris presents two specific 2a2 and 44 pairs andH. annuus a specific 33 pair. InH. argophyllus 11 33 or 44 are never observed but are replaced by 13 and 31 pairs. Some globulins, poorly represented, are of forms but present chains of higher molecular weights (in the range 54–56 kDa). Expressing variations in the banding patterns between these species by the use of a similarity index reveals complete identity between the three populations ofH. annuus. Identity between the twoH. petiolaris studied is also observed.H. annuus andH. argophyllus appear to be closer to each other thanH. petiolaris concerning the seed storage globulins. 相似文献
19.
A detailed investigation of hydrodynamic and conformational behavior has been made of the HM-crystallin and -crystallins of bovine lens. Results from this study indicated that HM (high-molecular-weight -crystallin) and (low-molecular-weight -crystallin) possess considerable size and charge heterogeneities in their native structures and subunit polypeptides, respectively. Sedimentation velocity showed a heterogeneous polydisperse system of HM with an average sedimentation coefficient of about 50 S and a more homogeneous system of -crystallin of 20 S. Viscosity and circular dichroism studies pointed to a compact and globular shape of dominant -sheet conformation for -crystallin, yet a highly asymmetrical and aggregated form for HM. The conformational stability of -crystallin was investigated in the presence of various denaturants. The evidence presented shows that hydrogen bonding is the main force in maintaining the quaternary structure of compact native -crystallin. Conformational flexibility of -crystallin demonstrated in the equilibrium unfolding study indicated a multistep transition that made the extraction of thermodynamic data from the heat denaturation study difficult. Temperature perturbation on -crystallin suggested the possible involvement of hydrophobic interaction in the aggregation process, leading to the formation of HM from -crystallin. The comparison of conformational properties between HM and -crystallin strongly indicated that HM is a denatured form of -crystallin. 相似文献
20.
Rudolf Geyer Silvia Diabaté Hildegard Geyer Hans-Dieter Klenk Heiner Niemann Stephan Stirm 《Glycoconjugate journal》1987,4(1):17-32
Fowl plague virus, strain Dutch, was metabolically labeled withd-[2-3H]mannose, or withd-[6-3H]glucosamine, and the small subunit (HA2; 0.8 mg in total) of the viral hemagglutinin was isolated by preparative sodium dodecylsulfate-polyacrylamide gel electrophoresis. After proteolytic digestion, the radioactive oligosaccharides were sequentially liberated from the glycopeptides by treatment with different endo--N-acetylglucosaminidases and with peptide:N-glycosidase or, finally, by hydrazinolysis. In this manner, four groups of glycans could be obtained by consecutive gel filtrations and were subfractionated by HPLC. The structures of the individual oligosaccharides were analyzed by micromethylation, by acetolysis or by digestion with exoglycosidases. The major species amongst the high mannose glycans at Ans-406 of the viral glycopolypeptide were found to be Man1-2Man1-3(Man1-2Man1-6)Man1-6(Man1-2Man1-2Man1-3)Man1-4GlcNac1-4GlcNAc and Man1-3(Man1-2Man1-6)Man1-6(Man1-2Man1-2Man1-3)Man1-4GlcNAc1-4GlcNAc, while the complex glycans at Asn-478 are predominantly GlcNAc1-2Man1-3(GlcNAc1-2Man1-6)Man1-4GlcNAc1-4GlcNAc (lacking, in part, one of the outerN-acetylglucosamine residues) and GlcNAc1-2Man1-3(Gal1-4GlcNAc1-2Man1-6)Man1-4GlcNAc1-4GlcNAc.Abbreviation BSA
bovine serum albumin
- endo D (F,H)
endo--N-acetyl-d-glucosaminidase D (F,H)
- HA
hemagglutinin (HA1, large subunit of HA
- HA2
small subunit
- FPV
fowl plague virus
- PNGase F
peptide:N-glycosidase F
- SDS
sodium dodecylsulfate 相似文献