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1.
Anna Marina Liberati Michael Horisberger Monica Schippa Francesco Di Clemente Marco Fizzotti Stefano Filippo Maria Grazia Proietti Sergio Arzano Paolo Berruto Lucia Palmisano Saverio Cinieri 《Cancer immunology, immunotherapy : CII》1991,34(2):115-122
Summary Ten hairy-cell leukemia patients were treated with interferon (IFN-) at a dose rate of 2 × 106 IU/m2 × 5 days for 4 weeks (induction therapy) and, thereafter, at the same dose three times a week for 11 months (maintenance therapy). The effect of this treatment on serum neopterin, 2-microglobulin, (2–5)oligoadenylate [(2–5)A
n
] levels, intracellular (2–5)A
n
values and human Mx protein synthesis was analysed. There were significant rises in serum neopterin and (2–5)A
n
levels during both induction and maintenance, whereas 2-microglobulin levels rose only during induction. Rises in intracellular (2–5)A
n
were documented mainly during induction, but they were not significantly higher than pretherapy values. IFN provoked an increase in human Mx protein synthesis over the entire induction — maintenance period, but was only significantly higher than baseline during induction. All markers proved useful for monitoring the effects of IFN dose schedules, but were not predictive of clinical outcome. Natural killer activity and IFN production, which were initially defective, followed a different trend from that of the other factors studied, in that increases were documented only late in the course of therapy when the disease was already in remission. 相似文献
2.
Comparative computer-assisted analysis was used to study putative GlpR regulons responsible for metabolism of glycerol and glycerol-3-phosphate in genomes of -, -, and -proteobacteria. New palindromic GlpR-binding signals were identified in -proteobacteria, consensus sequences being TGTTCGATAACGAACA for Enterobacteriaceae, wTTTTCGTATACGAAAAw for Pseudomonadaceae, and AATGCTCGATCGAGCATT for Vibrionaceae. The signals in - and -proteobacteria were also identified: they contained 3–4 direct TTTCGTT repeats separated by 3–4 nucleotide pairs. 相似文献
3.
2-Macroglobulin (2M) is a protease inhibitor that has separate binding sites for transforming growth factor- (TGF-) and -amyloid peptide (A), both of which have been identified in the 2M sequence. In the 3D-structure of 2M, TGF- occupies the 2M central cavity, overlapping with the space that can accommodate up to two molecules of protease. As a result, ternary 2M–protease complexes (2 mol protease/mol 2M) have been reported to not bind TGF-. The goal of the present study was to test whether binding of A to 2M is controlled by steric constraints imposed by associated proteases, similarly to TGF-. We confirmed that binary 2M–trypsin complex (1 mol trypsin/mol 2M) binds increased amounts of TGF-1, compared with native 2M, while ternary 2M–trypsin complex binds substantially decreased amounts of TGF-1. By contrast, A-binding to binary and ternary 2M–trypsin complex was equivalent. In both cases, binding was substantially increased compared with the negligible level observed with native 2M. Plasmin is a large protease (Mr ~82,000) that substantially occupies the 2M central cavity; however, 2M–plasmin complex also bound increased amounts of A, compared with native 2M. We conclude that A accesses its binding site, in 2M, from outside the 2M central cavity. The TGF--and A-binding sites are spatially separated not only in the primary sequence of 2M, but also in the 3D-structure. 相似文献
4.
The partition behaviour of -lactalbumin (la) and -lactoglobulin (lg) on PEG/(NH4)2SO4 system was studied. For purified proteins, a partition coefficient of 12.8 for la and 0.34 for lg, with mass recovery yields of 96.7% for la in the upper phase and 83.8% for lg in the lower phase was obtained, in 18% (w/w) PEG 900/14% (w/w) (NH4)2SO4 system, at pH 7. PEG/(NH4)2SO4 system was an economical alternative for the recovery and separation of the two proteins in cheese whey, allowing a 50% reduction in costs. An efficient and inexpensive separation of both proteins in cheese whey could be achieved, by using 16% (w/w) PEG 900/15% (w/w) (NH4)2SO4, at pH 7.5. 相似文献
5.
Deposition of amyloid peptide in human brain in the form of senile plaques is a neuropathological hallmark of Alzheimers disease (AD). Levels of a phospholipid breakdown product, glycerophosphocholine (GPC), also increase in AD brain. The effect of GPC on amyloid (1–40) peptide (A) aggregation in PBS buffer was investigated by circular dichroism and fluoresence spectroscopy; interactions of A and GPC with the intact erythrocyte membrane was examined by fluoresence spectroscopy. Fluorescamine labeled A studies indicate GPC enhances A aggregation. CD spectroscopy reveals that A in the presence of GPC adopts 14% more -sheet structure than does A alone. Fluorescamine anisotropy measurements show that GPC and A interact in the phospholipid head-group region of the erythrocyte membrane. In summary, both soluble A and GPC insert into the phospholipid head-group region of the membrane where they interact leading to -sheet formation in soluble A which enhances A aggregation. 相似文献
6.
Aeromonas salmonicida variants were characterized for alterations in their cell surface structure and used to examine reconstitution of the surface protein layer (A-layer). Variants lacking outer membrane O-polysaccharide were devoid of A-layer and excreted stainable floret-like material of the surface protein (A-protein). One variant, showing partial loss of O-polysaccharide, was associated with a disrupted A-layer and excretion of some A-protein. Variants lacking A-protein but possessing O-polysaccharide rapidly absorbed and concentrated sufficient excreted A-protein at the cell surface to coat the cells with a single confluent layer. Although differences in electrophoretic mobilities of A-proteins and O-polysaccharides from typical and atypical strains were evident, the different A-proteins and A-protein-deficient variants were interchangeable for reconstitution of a surface protein layer. No association of A-protein with cell surfaces of unrelated gram-negative bacteria was observed.Abbreviations A-layer
additional surface protein layer
- A-protein
surface protein
- Ast
Aeromonas salmonicida typical
- Asa
Aeromonas salmonicida atypical
- A-
phenotypically A-protein-negative variant
- O-
phenotypically O-polysaccharide-negative variant
- Owk
phenotypically O-polysaccharide weak variant
- BHI
brain heart infusion
- SDS-PAGE
sodium dodecylsulfate-polyacrylamide gel electrophoresis
- TEM
transmission electron microscopy 相似文献
7.
Barley -amylase was purified by ammonium sulfate fraction, ion-exchange, ultrafiltration, and gel filtration to homogeneity. The purified enzyme was partially digested with trypsin, and the reaction mixture was applied to a cyclohepta-amylose epoxy Sepharose 6B column. Bound fragments were eluted by free cyclohepta-amylose, lyophilized, and separated on Tricine gels. Four fragments were shown to interact with -cyclodextrin. The fragment that could be identified on the gel with the lowest molecular weight (11 kDa) was electroblotted onto PVDF membrane for sequencing. The N-terminal sequence of this fragment was determined with the N-terminal amino acid corresponding to Ala283 in the whole protein. The trypsin cleavage was at Lys282/Ala283 and the C-terminal cleavage occurred at Lys354/Ile355 to give a fragment size of 11 kDa as estimated by SDS-PAGE. The fragment would be located at the C-terminal region, forming a majority of the antiparallel -sheets in domain C and the 7-and 8-helices of the (/)8 domain. 相似文献
8.
-N-Acetyl-d-hexosaminidase from Aspergillus oryzae catalysed the stereo- and regiospecific formation of the 6-O-benzylated disaccharide derivatives GalNAc1-3(6- OBn)Gal-SEt and GlcNAc1-3(6-OBn)Gal-SEt, which were obtained in transglycosylation reactions employing ethyl 6- O-benzyl-1-thio--d-galactopyranoside as acceptor. Preparative amounts of the chitobiose derivative GlcNAc1- 3GlcNAc-OPhNO2-p was prepared as well. - N-Acetyl-d-hexosaminidase from bovine testes catalysed the specific synthesis of GlcNAc1-3(6-OBn)GlcNH2-SEt and GalNAc1-3(6-OBn)GlcNH2-SEt, employing ethyl 2-amino-6-O-benzyl-2-deoxy-1-thio--d-glucopyranoside as acceptor. -d-Glucuronidase from E. coli was found to catalyse the formation of GlcA1-3(6-OBn)GlcNH2- SEt employing the same acceptor. 相似文献
9.
In hippocampal neurons, 5-hydroxytryptamine (5-HT) activates an inwardly rectifying K+ current via G protein. We identified the K+ channel activated by 5-HT (K5-HT channel) and studied the effects of G protein subunits and nucleotides on the K+ channel kinetics in adult rat hippocampal neurons. In inside-out patches with 10 m 5-HT in the pipette, application of GTP (100 m) to the cytoplasmic side of the membrane activated an inwardly rectifying K+ channel with a slope conductance of 36±1 pS (symmetrical 140 mm K+) at –60 mV and a mean open time of 1.1±0.1 msec (n=5). Transducin activated the (K5-HT) channels and this was reversed by -GDP. Whether the K5-HT channel was activated endogenously (GTP, GTPS) or exogenously (), the presence of 1 mm ATP resulted in a 4-fold increase in channel activity due in large part to the prolongation of the open time duration. These effects of ATP were irreversible and not mimicked by AMPPMP, suggesting that phosphorylation might be involved. However, inhibitors of protein kinases A and C (H-7, staurosporine) and tyrosine kinase (tyrphostin 25) failed to block the effect of ATP. These results show that G activates the G protein-gated K+ channel in hippocampal neurons, and that ATP modifies the gating kinetics of the channel, resulting in increased open probability via as yet unknown pathways. 相似文献
10.
In Fuji, the production of ethylene was increased with the addition of AgNO3 and inhibited with the addition of 10 M aminoethoxyvinylglycine (AVG). The addition of 80 M AgNO3 to transformed explants of Fuji cultured on selection medium resulted in increased ethylene production (20 l l–1) at 3 weeks. Under examining the effect of AgNO3 in Fuji, the 40 M AgNO3 showed with higher 33.8% and 6.5% in the efficiency of regeneration and transformation. However, ethylene production in Gala explants treated with 10M AgNO3 (3 l l–1) decreased after 2 weeks compared with the control (5 l l–1). Although the regeneration efficiency of Gala with 10 M AgNO3 was higher (41.1%) than the control (20.1%), there was no significant difference in the transformation efficiency at the same concentration. Shoot regeneration of Fuji and Gala was completely inhibited with 10 M AVG. These results suggest that the addition of AgNO3 affects the efficiency of Agrobacterium-mediated gene transfer in Fuji.Eun Soo Seong, Ill Min Chung- These two Authors Contributed equally to this work 相似文献
11.
Richard P. Shank 《Neurochemical research》1992,17(5):423-430
Synaptosome-enriched preparations from rat and guinea pig brain tissue vigorously accumulated [3H]-adenosine 5-monophosphate ([3H]AMP). When the accumulation of [3H]AMP was determined using incubation periods of 30 s or less, high concentrations of adenosine, dipyridamole and soluflazine did not inhibit the accumulation of label appreciably. The accumulation of [3H]AMP was saturable, temperature-dependent, osmotic-sensitive and exhibited structural specificity. Based on the kinetics of uptake by different subcellular fractions, and the inhibitory effects of other nucleotides, the uptake of AMP appeared to be mediated by three saturable systems with Kt values of approximately 0.2, 6, and 100 M. The transport system with the highest affinity for AMP was selectively inhibited by guanosine 5-monophosphate, and its Vmax was several fold higher in a myelin-enriched fraction than in synaptosome-enriched fractions. The transport system with the Kt6 M was selectively inhibited by ,-methylene adenosine diphosphate, and its Vmax was several times higher in a fraction enriched in high-density synaptosomes than in fractions enriched in low-density synaptosomes or myelin. Both of these transport systems were potently inhibited by ATP and ADP. Nucleotides that were either weak or inactive as inhibitors of AMP transport included 3-AMP, cyclic AMP, guanosine 5-diphosphate, and the 5-mononucleotides of cytosine, inosine, and uridine. GTP consistently enhanced uptake at concentrations 1 M. The transport of AMP was not Na+-dependent and was not inhibited by membrane depolarization. This transport system may mediate the release of AMP for subsequent conversion to adenosine extracellularly.Abbreviations used HEPES
N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid
- NBTI
nitrobenzylthioinosine
- ,-MeADP
,-methylene adenosine diphosphate
- GTPgS
guanosine-5-O-(3-thiotriphosphate)
Special issue dedicated to Dr. Morris H. Aprison. 相似文献
12.
Suzuki Hisanori Buonamassa Daniela Tornese Weisz Alessandro 《Molecular and cellular biochemistry》1990,99(1):33-39
Summary The activity of poly (ADP-ribose) polymerase (ADPRP) and the content of 2,5-oligodenylates core (2,5An; n = 2,3 and 4) were measured in homogenates of the uterus and of the liver of immature rats immediately before (time 0) or at different times after injection of estradiol-valerate. ADPRP activity increased gradualy, starting 6 hours after estrogen injection, for about 4 days. Instead, the content of 2,5 An decreased by about 50% within 6 hours, and thereafter more slowly for 4 days to about 20% of starting values. Estrogen increased ADPRP activity and decreased 2,5An concentration also in the kidney and in the cardiac muscle of the same animals, but not in the skeletal muscle, where neither of the two parameters was affected. Injection of vehicle only (sesame oil) had no effect on ADPRP activity nor on 2,5An content of immature rat tissues. 相似文献
13.
DNA polymerases II () and III() are the only nuclear DNA polymerases known to possess an intrinsic 3 5 exonuclease in Saccharomyces cerevisiae. We have investigated the spontaneous mutator phenotypes of DNA polymerase and 3 5 exonuclease-deficient mutants, pol3-01 and pol2-4, respectively. pol3-01 and pol2-4 increased spontaneous mutation rates by factors of the order of 102 and 101, respectively, measured as URA3 forward mutation and his7-2 reversion. Surprisingly, a double mutant pol2-4 pol3-01 haploid was inviable. This was probably due to accumulation of unedited errors, since a pol2-4/pol2-4 pol3-01/pol3-01 diploid was viable, with the spontaneous his7-2 reversion rate increased by about 2 × 103-fold. Analysis of mutation rates of double mutants indicated that the 3 5 exonucleases of DNA polymerases and can act competitively and that, like the 3 5 exonuclease of DNA polymerase the 3 5 exonuclease of DNA polymerase acts in series with the PMS1 mismatch correction system. Mutational spectra at a URA3 gene placed in both orientations near to a defined replication origin provided evidence that the 3 5 exonucleases of DNA polymerases and act on opposite DNA strands, but were in sufficient to distinguish conclusively between different models of DNA replication. 相似文献
14.
Ahlert Schmidt 《Archives of microbiology》1977,112(3):263-270
Crude extracts of Rhodospirillum rubrum catalyzed the formation of acid-volatile radioactivity from (35S) sulfate, (35S) adenosine-5-phosphosulfate, and (35S) 3-phosphoadenosine-5-phosphosulfate. An enzyme fraction similar to APS-sulfotransferases from plant sources was purified 228-fold from Rhodospirillum rubrum. It is suggested here that this enzyme is specific for adenosine-5-phosphosulfate, because the purified enzyme fraction metabolized adenosine-5-phosphosulfate, however, only at a rate of 1/10 of that with adenosine-5-phosphosulfate. Further, the reaction with 3-phosphoadenosine-5-phosphosulfate was inhibited with 3-phosphoadenosine-5-phosphate whereas this nucleotide had no effect on the reaction with adenosine-5-phosphosulfate. For this activity with adenosine-5-phosphosulfate the name APS-sulfotransferase is suggested. This APS-sulfotransferase needs thiols for activity; good rates were obtained with either dithioerythritol or reduced glutathione; other thiols like cysteine, 2-3-dimercaptopropanol or mercaptoethanol are less effective. The electron donor methylviologen did not catalyze this reaction. The pH-optimum was about 9.0; the apparent K
m for adenosine-5-phosphosulfate was determined to be 0.05 mM with this so far purified enzyme fraction. Enzyme activity was increased with K2SO4 and Na2SO4 and was inhibited by 5-AMP. These properties are similar to assimilatory APS-sulfotransferases from spinach and Chlorella.Abbreviations APS
adenosine-5-phosphosulfate
- PAPS
3-phosphoadenosine-5-phosphosulfate
- 5-AMP
adenosine-5-monophosphate
- 3-AMP
adenosine-3-monophosphate
- 3-5-ADP
3-phosphoadenosine-5-phosphate (PAP)
- DTE
dithiorythritol
- GSH
reduced glutathione
- BAL
2-3-dimercaptopropanol 相似文献
15.
Amyloid beta peptide (A) is a small peptide present in normal cells and aggregated A is the main constituent of the extracellular amyloid plaques found in Alzheimers disease (AD) brain. Recent studies suggest that soluble A oligomers are neurotoxic rather than amyloid fibrils found in amyloid plaques. This study using multidimensional NMR spectroscopy and circular dichroism (CD) provides the first direct evidence that alterations in membrane structure can trigger the conversion of soluble -helical monomeric A into oligomeric A in a -sheet conformation. 相似文献
16.
Previous studies from this laboratory have demonstrated unambiguously that the isolated chain of human adult hemoglobin binds human haptoglobin (Hp). In the present work, the ability of the isolated subunits of haptoglobin and hemoglobin to form complexes is investigated. In quantitative radiometric adsorbent titrations, the H chain of haptoglobin bound to hemoglobin whereas the L chain had no binding activity. Also, the H chain of haptoglobin bound to the isolated and subunits of hemoglobin, but its binding to the or chain was less than the binding it exhibits to hemoglobin. The isolated L chain was able to reassociate with the H chain to form a complex that binds to hemoglobin or its subunits. Although the L chains had no binding activity, its association with the H chain increased the binding of the latter to Hb or its isolated and subunits suggesting a more indirect role for the L chain in haptoglobin-hemoglobin interactions. 相似文献
17.
Effects of continuous pentobarbital administration on binding characteristics of [3H]muscimol were examined by autoradiography, and levels of GABAA receptor 2-subunit mRNA were investigated by in situ hybridization histochemistry in the rat brain. In order to eliminate the induction of hepatic metabolism by systemic administration of pentobarbital, an i.c.v. infusion model of tolerance to and withdrawal from pentobarbital was used. An experimental model of barbiturate tolerance and withdrawal was developed using i.c.v. infusion of pentobarbital (300 g/10 l/hr for 7 days) by osmotic minipumps and abrupt withdrawal from pentobarbital. The levels of [3H]muscimol binding were elevated in cingulate of frontal cortex (46%) and granule layer of cerebellum (32%) of rats 24-hr after withdrawal from pentobarbital, while it was only elevated in cingulate (58%) of tolerant rats. The GABAA receptor 2-subunit mRNA was increased in the withdrawal rats only: in the cortex (9–14%), hippocampus (15–21%), inferior colliculus (21%), and granule layer of cerebellum (24%). These results show the involvement of GABAA receptor and its 2-subunit up-regulations in pentobarbital withdrawal rats, and suggest that the levels of [3H]muscimol binding and GABAA receptor 2-subunit mRNA are altered in a region-specific manner during pentobarbital withdrawal. 相似文献
18.
Summary When adenosine cyclic 2,3-phosphate is evaporated from solution in the presence of simple catalysts such as aliphatic diamines at alkaline pH, and maintained in a dry state at moderate temperatures (25-85°C), self-polymerization to give oligonucleotides of chainlength up to at least 6 is observed. The products contain an excess of [35]-linkages over [25]-linkages. The effects of different catalysts and reaction conditions on the efficiency of the reaction are described. The prebiological relevance of these reactions is discussed. 相似文献
19.
Alternating poly(Glu-Leu) exhibits a random coilstructure in pure water at neutral pH. The addition of 0.5 equiv of Ca2+ induces a coil-to--sheet transitionand the addition of 0.15 equivof Fe3+ induces a coil-to--helixtransition. Conformational competition between thesetwo structures was studied by mixing preformed-sheets and -helices in differentproportions. Circular dichroism spectra clearly show that-sheets are favored at the expense of-helices in -sheet-rich mixtures. 相似文献
20.
Rajendra Prasad Roy Parimala Nacharaju Ronald L. Nagel A. Seetharama Acharya 《Journal of Protein Chemistry》1995,14(2):81-88
Interspecies hybrids of HbA and Hb from mouse C57BL/10 [
2
M
2
H
and
2
H
2
M
(H=human, M=mouse)], representing 19 and 27 sequence differences per dimers (as compared with human dimer) have been generatedin vitro. The efficiency of the assembly of the interspecies hybrids by the alloplex intermediate pathway is about twofold higher than the low-pH-mediated subunit approach. The interspecies hybrids exhibit a cooperative O2 binding. The intrinsic O2 affinity of mouse Hb is slightly lower than HbA, while the 2,3-diphosphoglycerate (DPG) effect is comparable. Interestingly, the interspecies hybrid
2
M
2
H
has high O2 affinity (compared to either human or mouse Hb), while the interspecies hybrid
2
H
2
M
exhibits a very low O2 affinity. These results suggest that the mouse chain generates a tetramer with very low oxygen affinity. However, the complementarity of the mouse and chains generates a set of unique interactions that compensate for the low-oxygen-affinity propensity of the mouse chain. DPG binds the tetramer in the central cavity formed by the two subunits, hence the DPG effects on the interspecies hybrids should be as in the parent molecule. However, the results of the present study demonstrate that the DPG binding pocket is influenced by the nature of the chain present in the tetramer. The mouse chain reduces considerably the DPG right shift of the O2 affinity of the human-chain containing hybrid. Sequence analysis suggest that perturbations of the
1
1 (not the
1
2) are communicated to the DPG binding pocket in the presence of the alien subunit, and are the primary determinant of the ligand binding properties. The results have implications for the design of Hb-based blood substitutes and understanding of the inhibitory potential of mouse chains in transgenic mouse expressing human
S chains. 相似文献