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21.
The multicatalytic proteinase from rat skeletal muscle, a non-lysosomal high molecular weight enzyme active at neutral to alkaline pH, has been examined in the electron microscope as well as by dynamic laser light scattering. Both methods reveal monodisperse particles. Electron micrographs show a cylinder-shaped complex with a diameter of 11 nm and a length of 16 nm in negatively stained, and a diameter of 9.6 nm and a length of 14.3 nm in freeze-dried, heavy metal replicated specimens. The molecule is composed of four rings or disks. 相似文献
22.
C A Mikoryak B W Elliott M E Kimball L A Steiner 《Journal of immunology (Baltimore, Md. : 1950)》1986,136(1):217-223
The immunoglobulins of the bullfrog Rana catesbeiana are unusual in that, in all classes, the light chains are not disulfide bonded to heavy chains or to other light chains. Moreover, the light chains contain six, rather than the usual five, residues of half-cystine. As none of these half-cystines is in the sulfhydryl form or is alkylated after mild reduction, we suggested that the light chains probably contain three intrachain disulfide bridges. We have now carried out experiments to confirm the existence of an extra intrachain disulfide bridge in Rana catesbeiana light chains and to determine its location. Disulfide bridge assignments were based on 1) isolation and sequence analysis of S-(carboxymethyl)cysteine-containing peptides and 2) isolation, from unreduced light chains, of peptides containing a disulfide bridge. Half-cystine residues were found at positions 134 and 194, and these were shown to be joined in the conserved intradomain disulfide bridge. In addition, we found that a residue of half-cystine, located at the third position from the carboxy-terminus, forms a disulfide bridge with a half-cystine at position 119, near the amino-terminus of the domain, the latter residue replacing a proline that has been found at this position in all other light chains. An intrachain disulfide bridge has not been found at this location in any other light chain. 相似文献
23.
G Steiner E Tschachler M Tani T R Malek E M Shevach W Holter W Knapp K Wolff G Stingl 《Journal of immunology (Baltimore, Md. : 1950)》1986,137(1):155-159
Rat monoclonal antibodies 3C7 and 7D4 detect two distinct functional regions of the murine interleukin 2 (IL 2) receptor. When studying the emergence kinetics of IL 2 receptors in mixed epidermal cell (EC)-lymphocyte cultures by using 3C7 and 7D4 in an indirect immunofluorescence assay, we regularly encountered a distinctive membrane fluorescence not only on lymphocytes, but also on a subpopulation of cells exhibiting a dendritic morphology. Reasoning that these 3C7/7D4-reactive dendritic cells might represent a subpopulation of epidermal dendritic cells, we studied mouse EC for the presence of 3C7/7D4- reactive cells. Although 3C7/7D4 reactivity was never detected on freshly isolated EC or on epidermal sheets, a small number of 3C7/7D4+ cells was encountered after 24 to 48 hr of culture. These cells exhibited a dendritic shape, expressed Ia antigens, lacked Thy-1 antigens, and displayed the ultrastructural features of Langerhans cells (LC) with the notable exception of Birbeck granules. Although after 24 hr, only 20% of Ia+ EC were 3C7/7D4+, the vast majority of LC displayed 3C7/7D4 binding sites after 4 to 5 days of culture. Preincubation of cultured LC-enriched EC with recombinant human IL 2 prevented subsequent 3C7-but not 7D4-binding to these cells. Western blot analysis of 7D4-reactive material of detergent extracts from LC-enriched EC revealed three bands in the same m.w. range as reported for CTLL cells. These results demonstrate that cultured LC express IL 2 receptors and may bear important implications for a better understanding of growth regulation, differentiation, and immunologic functions of LC. 相似文献
24.
Johannes Pohlner Thomas F. Meyer Paul A. Manning 《Molecular & general genetics : MGG》1986,205(3):501-506
Summary Fusion proteins comprising the amino-terminal 99 amino acids of the bacteriophage MS2 replicase and various portions of OmpV a major outer membrane protein of Vibrio cholerae were expressed in Escherichia coli K12. These fusions were expressed under the control of the PL promoter of bacteriophage , and expression was controlled using a cIts repressor. Fusions occurring within the secretory signal sequence of OmpV gave rise to the production of mature OmpV. The efficiency, however, decreased with progressive deletion of the signal sequence within the fusions. The reactivity of various OmpV fusions with antisera raised against purified OmpV and whole bacteria demonstrated the existence of two antigenic domains: one present in the denatured form and another in the membrane-associated form of OmpV. These domains correspond to markedly hydrophilic regions of the protein as would be predicted for surface-exposed epitopes. 相似文献
25.
The transport of several amino acids with different side-chain characteristics has been investigated in the aquatic liverwort Riccia fluitans. i) The saturation of system I (neutral amino acids) by addition of excess -aminoisobutyric acid to the external medium completely eliminated the electrical effects which are usually set off by neutral amino acids. Under these conditions arginine and lysine significantly depolarized the plasmalemma. ii) L- and D-lysine/arginine were discriminated against in favour of the L-isomers. iii) Increasing the external proton concentration in the interval pH 9 to 4.5 stimulated plasmalemma depolarization, electrical net current, and uptake of [14C]-basic amino acids. iv) Uptake of [14C]-glutamic acid took place only at acidic pHs. v) [14C]-histidine uptake had an optimum between pH 6 and 5.5. vi) Overlapping of the transport of basic, neutral, and acidic amino acids was common. It is suggested that besides system I, a second system (II), specific for basic amino acids, exists in the plasmalemma of Riccia fluitans. It is concluded that the amino-acid molecule with an uncharged side chain is the substrate for system I, which also binds and transports the neutral species of acidic amino acids, whereas system II is specific for amino acids with a positively charged side chain. The possibility of system II being a proton cotransport is discussed.Abbreviation AiB
-aminoisobutyric acid 相似文献
26.
The photocycle of the chloride pump halorhodopsin. I: Azide-catalyzed deprotonation of the chromophore is a side reaction of photocycle intermediates inactivating the pump 下载免费PDF全文
Halorhodopsin, the light-driven chloride pump of halobacteria, undergoes a photochemical cycle in the 10 ms range. Two intermediates, HR640 and HR520, accumulate in the photosteady state after short times (within 100 ms) of illumination. Upon prolonged illumination a third species, HRL410 accumulates, which is formed from HR520/HR640 by deprotonation of the chromophore in a side reaction of the photocycle. In the dark, HRL410 requires several minutes to reconvert thermally to HR478. Thus, molecules in the HRL410 state must be inactive pumps since their maximal turnover number could only be a few per hour. Inorganic bases, such as azide, catalyze the deprotonation of HR520/HR640 as well as the reprotonation of HRL410. Both reactions are accelerated several hundred times by azide but the photosteady-state concentration of HRL410 remains unchanged. 相似文献
27.
Dr. Gerard J. McGarrity Lindsay Gamon Theodor Steiner Joseph Tully Hitoshi Kotani 《Current microbiology》1985,12(2):107-112
Uridine phosphorylase activity has been used to detect mycoplasmas in cell cultures by measuring formation of14C-uracil from14C-uridine. In this report we show that all species ofMycoplasma, Acholeplasma, andUreaplasma tested exhibited uridine phorphorylase activity. Among the genusSpiroplasma, serogroups I-1, I-3, I-5, I-7, I-8, IV, XIII, and XIV lacked uridine phosphorylase activity.Present address: Ciba-Geigy, Basel, Switzerland. 相似文献
28.
Conformational alterations occurring in bovine alpha 2-macroglobulin (alpha 2M) resulting from proteolysis and nucleophilic modification have been monitored by UV difference spectra, circular dichroism, and changes in the fluorescence of 6-(p-toluidino)-2-naphthalenesulfonate (TNS) and bis(8-anilino-1-naphthalenesulfonate) (Bis-ANS). The results of this study indicate that these two dyes appear capable of differentiating between conformational changes induced by proteolysis and those induced by methylamine treatment. It appears that TNS is a sensitive probe for monitoring protease-induced but not methylamine-induced conformational changes in bovine alpha 2M. Bis-ANS, on the other hand, appears suitable for monitoring conformational changes induced by methylamine treatment or proteolysis of the molecule and was used as a probe to monitor the kinetics of the conformational change induced by methylamine treatment. It was found that the conformational change did not occur simultaneously with cleavage of the thiol ester bonds by the nucleophile, measured by titration of free sulfhydryl groups with 5,5'-dithiobis(2-nitrobenzoate). The data are consistent with a model in which initial nucleophilic attack results in exposure of sulfhydryl groups, resulting in a conformational change measured by an increase in fluorescence. This event is followed by a unimolecular step representing a conformational change in the protein that results in a further increase in the fluorescence signal. The second-order rate constant for hydrolysis of the thiol ester bonds was determined to be 3.4 +/- 1.0 M-1 s-1, while the rate constant for the conformational change was (4.4 +/- 0.8) X 10(-4) s-1. 相似文献
29.
Lack of effect of somatostatin on the stimulation of hepatic glycogenolysis by epinephrine in isolated canine hepatocytes 总被引:1,自引:0,他引:1
R W Stevenson K E Steiner D R Green A D Cherrington 《Biochimica et biophysica acta》1984,804(4):414-418
The effects of somatostatin on epinephrine's ability to stimulate glucose output have been examined in hepatocytes isolated from dogs fasted overnight. Half-maximal stimulation of phosphorylase a activity and glucose output occurred at an epinephrine concentration of approx. 5 X 10(-9) M. Somatostatin at 10, 100 or 1000 ng/ml had no effect on the ability of a maximal (1 X 10(-7) M) and a submaximal (1 X 10(-8) M) dose of epinephrine to activate phosphorylase at 2 min, or to stimulate glucose output over 20 min. Since the doses of somatostatin used in the present study are up to 50-fold higher than the blood concentrations commonly found when somatostatin is used in vivo to inhibit pancreatic hormone secretion, it seems unlikely that use of somatostatin in this way would affect stimulation of hepatic glycogenolysis by epinephrine in vivo. 相似文献
30.
Halide binding by the purified halorhodopsin chromoprotein. I. Effects on the chromophore 总被引:4,自引:0,他引:4
The halorhodopsin chromoprotein, a retinal-protein complex with an apparent molecular mass of 20 kilo-daltons, exhibits all of the halide-dependent effects found for the chromophore of functional halorhodopsin in cell envelope vesicles. With increasing halide concentration (a) an alkali-dependent 580/410 nm chromophore equilibrium (attributed to reversible deprotonation of the retinal Schiff's base) is shifted toward the 580-nm chromophore and (b) the flash-induced photocycle proceeds increasingly via P520, rather than via P660. The halide-binding site(s) responsible for these effects must reside, therefore, in the chromoprotein. Chloride and bromide are about equivalent, but iodide is much less effective in these effects and in being transported. Several other anions, i.e. thiocyanate, nitrate, phosphate, and acetate, affect the absorption maximum of the chromophore but do not allow the production of P520 upon flash illumination and are not transported. However, these ions appear to compete with chloride in the flash experiments. These observations suggest that binding of anions to a relatively nonspecific site affects the protonation state of the Schiff's base in the chromophore. Either this site directly or a more specific site, connected to the first one by a sequential pathway, is involved with the photocycle intermediates and with chloride transport by halorhodopsin. 相似文献