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11.
Helmut Tr?ster Wolfgang Kissel Michael F. Trendelenburg Ansgar Hofmann 《Molecular & general genetics : MGG》1989,217(2-3):533-535
Summary Eight representative recombinant background clones of λEMBL3 were analysed usingKpnI,BamHI,SalI,EcoRI andHindIII digestion. We found that λEMBL3 carries its own left arm in theBamHI cloning site. In this way, recombinant molecules were found to be generated which can grow onEscherichia coli strain NM539. In all cases analysed, the left arm DNA was inserted in a head to tail orientation. Seven clones carried a
restoredBamHI site at thecos site-BamHI site connection. In the region where the inserted left arm and the right arm were ligated,BamHI cloning produces a large palindromic sequence consisting of two polylinkers. ThisBamHI site was incompletely cleaved in all cases analysed. We assume that a part of the λ DNA molecule in this region shows a
cruciform structure prohibiting recognition or cleavage of this site by restriction endonucleaseBamHI. 相似文献
12.
Helmut Sigel 《Biological trace element research》1989,21(1):49-59
The self-association of nucleosides decreases within the series adenosine>guanosine>inosine>cytidine ≈uridine. The same trend is observed for the corresponding nucleotides, though less pronounced, as the charge effect governs series like adenosine ? AMP2?>ADP3??ATP4?. Protonation of adenosine considerably reduces its self-stacking tendency: this is different with ATP4?, where a maximum is reached for H2(ATP)2? caused by additional ionic interactions in the [H2(ATP)]2 4? dimer. Metal ion coordination may promote self-association, e.g., of ATP4? via (mainly) charge neutralization (Mg2+) and the formation of intermolecular bridges in dimeric stacks (Zn2+, Cd2+). These results allow definition of conditions with negligible self-association and thus the determination of the stability and structure of monomeric nucleotide complexes in aqueous solution, e.g., quantification of macrochelate formation in M(ATP)2? complexes. Some biological implications of the results are indicated. 相似文献
13.
Properties of several protein kinases that copurify with rat spinal cord neurofilaments 总被引:2,自引:0,他引:2
C B Caputo L A Sygowski W F Brunner C W Scott A I Salama 《Biochimica et biophysica acta》1989,1012(3):299-307
Several protein kinases that copurify with neurofilaments (NF) were identified and each kinase was assessed for its ability to phosphorylate NF proteins. NFs were isolated using an axonal flotation procedure and the kinases were extracted from NFs with 0.8 M KCl. NF kinases were incubated with peptide substrates for selected protein kinases, [32P]ATP and protein kinase cofactors and inhibitors to characterize the kinases. Using peptide substrates, three types of kinase were identified, and a fourth was identified using NF protein as substrate. The first three kinases were the catalytic subunit of cAMP-dependent protein kinase, calcium-calmodulin dependent protein kinase II and a cofactor-independent kinase that phosphorylated prepro VIP sequence 156-170 and was inhibited by heparin. Using NF proteins as substrate, a fourth kinase was identified which was cofactor-independent and was not inhibited by heparin. Neither cofactor-independent kinase was casein kinase II. NF proteins were phosphorylated in vitro on serine and threonine, primarily by the two cofactor-independent kinases. Using [alpha-32P]8-N3ATP for affinity labeling, one kinase of 43,800 Da was identified. Thus, in addition to cAMP-dependent protein kinase and calcium-calmodulin dependent protein kinase II, two kinases have been found which are primarily responsible for NF phosphorylation in vitro and are cofactor-independent. 相似文献
14.
Helmut Krüger Werner Schr?der Klaus Buchner Ferdinand Hucho 《Journal of Protein Chemistry》1990,9(4):467-473
Inhibition of protein kinase C (PKC) by calmodulin is investigated and we describe the localization of inhibitory sequences within the calmodulin molecule. We present evidence that calmodulin inhibits PKC through an inhibition of the activation of PKC associated with lipid membranes: Binding of PKC to lipid vesicles is not affected, but activation is abolished. The potent calmodulin antagonist R24571 (calmidazol) did not affect the inhibition of PKC by calmodulin at concentrations up to 10–5 M. Two tryptic fragments of calmodulin were isolated which inhibited PKC. They were only slightly less potent than intact calmodulin with an IC50 of 6 µ M compared to 1 µ M of intact calmodulin. They were identified as Ser38-Arg74 and His107-Lys148. Each of the inhibiting fragments contains an intact Ca2+-binding domain with complete helix-loop-helix structure (EF hand). Other calmodulin peptides showed only weak inhibitory activity. Both fragments did not stimulate cAMP phosphodiesterase even at concentrations 100-fold higher than the calmodulin concentration needed for maximal stimulation. None of the fragments acted as a calmodulin antagonist. 相似文献
15.
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17.
Crude extracts or supernatants of broken cells of Clostridium formicoaceticum reduce unbranched, branched, saturated and unsaturated carboxylates at the expense of carbon monoxide to the corresponding alcohols. The presence of viologens with redox potentials varying from E
0=-295 to-650 mV decreased the rate of propionate reduction. The more the propionate reduction was diminished the more formate was formed from carbon monoxide. The lowest propionate reduction and highest formate formation was observed with methylviologen. The carbon-carbon double bond of E-2-methyl-butenoate was only hydrogenated when a viologen was present. Formate as electron donor led only in the presence of viologens to the formation of propanol from propionate. The reduction of propionate at the expense of a reduced viologen can be followed in cuvettes. With respect to propionate Michaelis Menten behavior was observed. Experiments are described which lead to the assumption that the carboxylates are reduced in a non-activated form. That would be new type of biological reduction.Non-standard abbreviations glc
Gas liquid chromatography
- HPLC
high performance liquid chromatography
- RP
reverse phase; Mediators (the figures in parenthesis of the mediators are redox potentials E
0 in mV)
- CAV2+
carbamoylmethylviologen, 1,1-carbamoyl-4,4-dipyridinium dication (E
0=-296 mV)
- BV2+
benzylviologen, 1,1-dibenzyl-4,4-dipyridinium dication (E
0=-360 mV)
- MV
methylviologen, 1,1-dimethyl-4,4-dipyridinium-dication (E
0=-444 mV)
- DMDQ2+
dimethyldiquat, 4,4-dimethyl-2,2-dipyridino-1,1-ethylendication (E
0=-514 mV)
- TMV2+
tetramethylviologen, 1,1,4,4-tetramethyl-4,4-dipyridinium dication (E
0=-550 mV)
- PDQ2+
propyldiquat, 2,2-dipyridino-1,1-propenyl dication (E
0=-550 mV)
- DMPDQ2+
dimethylpropyldiquat, 4,4-dimethyl-2,2-dipyridino-1,1-propenyl dication (E
0=-656 mV)
- PN
productivity number=mmol product (obtained by the uptake of one pair of electrons) x (biocatalyst (dry weight) kg)-1×h-1 相似文献
18.
Helmut Segner Roland Marthaler Michael Linnenbach 《Environmental Biology of Fishes》1988,21(2):153-159
Synopsis Young (7–10 days after hatching) brown trout (Salmo trutta) exposed for 5 days to pH 5 in high calcium water and at 2 temperatures (12°, 4°C) in the laboratory displayed no alterations
in growth or in mucous cell concentration and volume, compared to the control group kept at pH 7.2. Contamination of acid-stressed
young with 230 μg All-1 resulted in significant growth depression and Al accumulation, but in no changes of mucous cell morphometrics. Field tests
in low calcium water produced high mortality at low pH (5.1), but showed consistent effects on mucous cells as in laboratory
experiments. Three-month-old juveniles of brown trout, subjected to decreased pH values at 12° and in high calcium water for
8 days exhibited mucous cell hyperplasia (without hypertrophy) within 3 h of the acid addition. After 120 h sloughing of the
integument occurred with full recovery not possible within a 4-day-recovery period. Although the results presently apply only
to hard water conditions, the differences between juveniles and recently hatched young in tolerance to pH- and Al-mediated
stress may also be of importance for soft waters affected by acid rain. 相似文献
19.
20.
Tetanus Toxin and Botulinum A and C Neurotoxins Inhibit Noradrenaline Release from Cultured Mouse Brain 总被引:4,自引:3,他引:1
The specific binding protein for substance P (SP) was solubilized in an active form from the crude mitochondrial (P2) fraction of bovine brainstem. After incubation with 3-[(3-cholamidopropyl)dimethylammonio]-1-propane sulfonate (CHAPS) and 0.1 M NaCl at 0 degrees C for 30 min, the SP binding to the supernatant fraction (100,000 g, 60 min) was determined by the glass fiber filtration method reported by Bruns et al. (1983). The specific [3H]SP binding to the solubilized fraction was highly specific for SP and was displaced by nanomolar concentrations of SP and physalaemin, but only by micromolar concentrations of eledoisin. In addition, the binding was inhibited by GTP (approximately 40% of the specific binding decreased by 10 microM GTP) in both preparations. These results were virtually identical to those of P2 membrane preparations and suggested that this high-affinity SP binding site belongs to the SP-P type. Scatchard analyses of SP binding to the solubilized fraction revealed a single saturable component with a Bmax of 22.0 +/- 5.10 fmol/mg protein and a KD of 0.79 nM, and these values are almost the same as those obtained in the P2 fraction (Bmax = 31.3 +/- 3.56 fmol/mg protein, KD = 0.82 nM). Gel filtration analysis showed that the detergent-SP binding protein complex has two calculated molecular weights of greater than 1,000,000 and 55,000-60,000 (a corresponding Stokes radius of 35.5 nm). 相似文献