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131.
Structural studies of the lysozyme coded by the pneumococcal phage Cp-1. Conformational changes induced by choline 总被引:2,自引:0,他引:2
The CPL-1 lysozyme coded by the pneumococcal phage Cp-1 has been overproduced in Escherichia coli under the control of a modified lipoprotein lactose promoter. This result has provided the conditions to analyse the CPL-1 secondary structure by circular dichroism (CD). The CD spectra recorded in the far-ultraviolet region showed, at neutral pH, two minima at 210 nm and 230 nm and a shoulder at 217 nm, whereas two bands at 260 nm and 295 nm were observed in the near-ultraviolet region. It has been estimated, by using the CDPROT program, that the protein is composed of 19% alpha-helix, 32% beta-sheet, 28% beta-turn and 21% random coil. Minor changes in the CD spectra were detected either when the pH was varied over 6-10 or when the ionic strength was increased to 1 M NaCl. Choline, a well known modulator of the enzyme activity that is present in the pneumococcal cell wall, induced remarkable changes in the intensities of the bands at 210, 230 and 295 nm, with the appearance of an unusual positive band at 225 nm. The conformational change was reversible and correlated with the competitive inhibitory effect of choline on the lysozyme activity, supporting, by a new and direct experimental approach, the basic role of choline in the recognition of the cell wall substrate. The analyses of the secondary structure prediction and the CD data reported here are compatible with the two-domain structure of CPL-1 reinforce our hypothesis that the C-terminal region is directly involved in the binding of the enzyme to the pneumococcal teichoic and lipoteichoic acids. 相似文献
132.
Sequence of the Streptococcus pneumoniae bacteriophage HB-3 amidase reveals high homology with the major host autolysin. 总被引:10,自引:5,他引:5
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We have sequenced a DNA fragment containing the pneumococcal bacteriophage HB-3 hbl gene, which codes for the phage lytic amidase. A remarkable nucleotide similarity (87.1%) between the lytA gene, coding for the pneumococcal amidase, the major autolysin of Streptococcus pneumoniae, and the hbl gene was found. This similarity completely disappeared outside the open reading frames coding for both amidases. The hbl gene transformed amidase-deficient strains of S. pneumoniae to the wild-type phenotype, and Southern blotting experiments provided evidence for recombination between donor and recipient genes. A comprehensive evaluation of these and previous results on the peptidoglycan hydrolases of S. pneumoniae and its bacteriophages suggested that recombination mechanisms participate in the evolution of the genes coding for these enzymes. 相似文献
133.
Andrew R. Cross Owen T. G. Jones Rudolfo Garcia Anthony W. Segal 《The Biochemical journal》1982,208(3):759-763
A plasma membrane fraction prepared from human neutrophils had a fluorescence resembling that of a fluorescent flavoprotein, with emission maximum near 520nm and excitation maxima near 380 and 460nm. The fluorescence emission and excitation properties of Triton N-101-solubilized membrane fraction resembled those of FAD. FAD was present in the membranes at a concentration of 417pmol/mg of protein and cytochrome b−245 at a concentration of 407pmol/mg of protein. In a 110-fold purified preparation of cytochrome b−245 the ratio of FAD:cytochrome b was 1:1. Analytical gradient centrifugation of neutrophil homogenates shows a coincidence of two cytochrome b peaks and two peaks of fluorescence, corresponding with plasma membrane and specific granule fractions; most of the FAD was non-fluorescent and located in fractions lighter than the plasma membrane. Plasma membrane fractions prepared from neutrophils of patients suffering from the X-linked form of chronic granulomatous disease lacked cytochrome b and contained 194pmol of FAD/mg of protein; plasma membrane fractions prepared from neutrophils of patients with the autosomal recessive form of chronic granulomatous disease contained both cytochrome b−245 and FAD in the normal range of concentrations in a ratio of 1:1. Phagocytic vesicles were prepared from normal neutrophils and found to contain FAD and cytochrome b in a ratio 2.22:1, suggesting that activation of neutrophils many involve the incorporation of an additional flavin into the membrane. Under anaerobic conditions in the presence of EDTA to act as an electron donor to a flavin, the cytochrome b−245 of neutrophil membranes was partly (12%) photoreducible, an effect increased to 100% by the addition of FMN. The extent of reduction of cytochrome b in an anaerobic neutrophil homogenate containing NADH increased from 30% to 70% on illumination. We suggest that these results indicate a close association between FAD and cytochrome b−245 and support a scheme for electron transport thus: [Formula: see text] 相似文献
134.
St. Ledoux J. Gutkowska R. Garcia G. Thibault M. Cantin J. Genest 《Histochemistry and cell biology》1982,74(3):329-339
Summary As a part of a microfluorometric investigation of the nucleoproteins of nuclei whose chromatin displays varying degrees of condensation, a comparison was made of mouse small thymocyte and hepatocyte nuclei stained with the acidic dye, brilliant sulfaflavine, at pH 2.8. These estimates of total protein content were compared with measurements obtained in similarly stained nuclei after extraction either with 0.4 N H2SO4 to remove all histones or with 0.35 M NaCl to remove nucleoplasmic proteins and some loosely bound non-histone chromosomal proteins. Treatment with 5% TCA at 60°C was used to remove nucleic acids and to reverse the effects of formaldehyde fixation. In all instances, the fluorescence of 2c hepatocyte nuclei greatly exceeded that of similarly treated thymocyte nuclei. While extraction with 0.4 N H2SO4 resulted in reductions of as much as 75% of the total fluorescence of small thymocyte nuclei, the losses of fluorescence in 2c hepatocyte nuclei amounted to only 20–30%. Nevertheless, the absolute values of fluorescence lost in both types of nuclei were very similar. After extraction in 0.35 M NaCl, thymocyte nuclei displayed slightly greater fluorescence than control thymocyte nuclei, while the total fluorescence of hepatocyte nuclei declined. In hepatocyte nuclei extracted with TCA, with and without treatment with 0.35 M NaCl, two populations of diploid nuclei were apparent: one corresponding to parenchymal cell nuclei and the other comprised of non-parenchymal cell nuclei. Only single diploid populations were visible in acid-extracted material. The ratios of 4c2c, 8c4c, and 8c2c hepatocyte nuclei in control, acid-extracted, and NaCl-extracted groups were generally lower than the expected 224 values. These results indicate that total nuclear histones may be estimated microfluorometrically by computing the difference between acid-extracted and unextracted preparations treated in otherwise equivalent ways. In addition, despite very similar absolute losses of fluorescence after removal of histones in thymocyte and 2c hepatocyte nuclei, the proportion of total protein ascribable to histones is much greater in thymocyte nuclei than in 2c hepatocyte nuclei — or, conversely, the percentage of total protein attributable to non-histone proteins is much greater in 2c hepatocyte nuclei than in thymocyte nuclei. 相似文献
135.
Suzanne Desjardins-Giasson Jolanta Gutkowska Raul Garcia Jacques Genest 《Prostaglandins & other lipid mediators》1982,24(1):105-114
Prostaglandin E2 (PGE2) and 6 keto-PGF1α, the stable metabolite of prostacyclin (PGI2), have been measured in the effluent of perfused rat mesenteric arteries by the use of a sensitive and specific radioimmunoadday (RIA) method. The PGE2 and 6-keto-PGF1α were continuousyl released by the unstimulated mesenteric artery over a period of 145 min. After 100 min of perfusion the release of PGE2 and 6-keto-PGF1α was 4.5 ± 8.4 pg/min and 254 ± 75 pg.min respectively, which is in accord with the general belief that PGI2 is the major PG synthesized by arterial tissue. Angiotensin II (AII) 5 ng/ml) induced an increased of PGE2 and 6-keto-PGF1α release without changing the perfusion pressure. The effect of norepinephrine (NE) injections on release of PGs depended on the duration of the stabilization period. The changes of perfusion pressure induced by NE were not related to changes in release of PGs. Thus, it seems that the increase of PG release induced by AII and NE was due to a direct effect of the drugs on the vascular wall. This may represent an important modulating mechanism in the regulation of vascular tone. 相似文献
136.
Carlo M. Ignoffo Clemente Garcia Michael J. Kroha 《Journal of invertebrate pathology》1982,39(2):198-202
The LD50 for larvae of Trichoplusia ni injected with blastospores of Nomuraea rileyi was 4.30 ± 1.16 hyphal bodies/larva; the LD50 for injected conidia was ca. 25,000 conidia/larva. The dose-mortality regression line for blastospores was Y = 4.6504 + 0.5487 X. Larval mortalities of Anticarsia gemmatalis and T. ni at 100 blastospores/larva were 0.4 ± 0.5% and 96.7 ± 1.9%, respectively. At a dosage of 25,000 conidia/larva, larval mortalities for A. gemmatalis and T. ni were 0.4 ± 0.5% and 43.1 ± 8.7%, respectively. Thus, larvae of A. gemmatalis were > 100 times and >200 times more resistant to injected conidia and blastospores, respectively, than were larvae of T. ni. Resistance of A. gemmatalis to N. rileyi may not be solely at the integumental barrier, as is often believed, but may also be a function of an internal physiological response. 相似文献
137.
To determine the usefulness of R-wave amplitude changes during exercise testing for the diagnosis of coronary artery disease (CAD) and to understand the discrepancies that have been described in the literature regarding their value, we studied two groups of patients by means of electrocardiographic (EKG) treadmill testing and coronary arteriography. Group I was composed of 149 patients who were studied prospectively. The specificity of R-wave changes measured from preexercise to immediately postexercise (SRV(5)) was 81%, but that of R-wave changes measured from preexercise to peak exercise (URV(5)) was 46%. A group of 156 patients (Group II) evaluated retrospectively showed a high specificity for the SRV(5) (84%) and poor specificity for the URV(5) (39%). The sensitivity of the SRV(5) was 38% in Group I and 42% in Group II. Therefore, if measured during the immediate postexercise period and not at peak exercise, changes in R-wave amplitude may be of value in the diagnosis of coronary artery disease by electrocardiographic exercise testing. 相似文献
138.
139.
140.
Acetone powders prepared from a 20,000g participate preparation from spinach leaf catalyzed several reactions involving monoacylglycerol and diacylglycerol. When these substrates were presented as Triton X-100-mixed micelles, diacylglycerol gave rise to free fatty acids, monoacylglycerol, triacylglycerols, and steryl esters, and in the presence of ethanol, small amounts of ethyl esters of fatty acid. Monoacylglycerol gave rise to free fatty acids and diacylglycerol, and in the presence of ethanol, large amounts of ethyl esters of fatty acid. In the presence of bovine serum albumin, the conversion of monoacylglycerol to free fatty acid was retarded. In the presence of bovine serum albumin, steryl ester was an important product from diacylglycerol. The system containing Triton X-100-mixed micelles and bovine serum albumin permitted analysis of reaction products which showed diacylglycerol to be an acyl donor in steryl ester biosynthesis. All reactions observed in the mixed micelle system were transacylation reactions involving various acceptors: dipalmitoylglycerol → monopalmitoylglycerol + palmitate; monopalmitoylglycerol → glycerol + palmitate; dipalmitoylglycerol + sterol → monopalmitoylglycerol + steryl palmitate; monopalmitoylglycerol + ethanol → ethyl palmitate + glycerol; monopalmitoylglycerol → dipalmitoylglycerol (+glycerol); dipalmitoylglycerol → tripalmitoylglycerol (+monopalmitoylglycerol). 相似文献