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The ability of peptides coded by the anti-sense strand of DNA to interact specifically with peptides coded by the sense strand has been evaluated. The sense peptide examined, ribonuclease S-peptide, was immobilized on a coated silica affinity chromatographic matrix. Anti-sense peptides were synthesized on the basis of the anti-sense DNA sequence for the S-peptide region in native pancreatic ribonuclease A. The interaction of synthetic anti-sense peptides with sense peptide was quantitated from the degree of retardation during chromatographic elution on the sense peptide affinity matrix in buffers with and without soluble competing sense peptide. Sense/anti-sense peptide interactions were found to occur with significant affinities with each of two anti-sense 20-residue peptides of opposite amino-to-carboxyl orientations and to weaken progressively with decreasing length of anti-sense peptide. The substantial chromatographic retardation of anti-sense peptides was specific, since it decreased as expected with increasing concentration of the soluble competing S-peptide, could not be mimicked by the elution of several control peptides (including S-peptide itself) on the S-peptide matrix, and did not occur with a blank chromatographic matrix (no S-peptide attached). The stoichiometry of anti-sense peptide binding to immobilized sense peptide was found to be far greater than 1:1, and at least 4-5:1, for the two 20-mer anti-sense peptides. In sum, the analytical affinity chromatographic experiments have established quantitatively that anti-sense peptide binding to sense peptides occurs in the ribonuclease S-peptide case and have identified some structural elements that govern these interactions.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
5.
Alexander E. Gad Carmi Lubitz-Omero Nurit Rosenberg Arie Altman 《Plant cell reports》1988,7(5):341-343
Negatively charged large unilamellar vesicles (LUV) were incubated with tobacco (Nicotiana tabacum var. xanthi) cell suspensions and with the cell-free medium of the cell suspensions. The extent of cell-LUV interaction was determined by the leakage of the LUV contents. Cells enhanced the leakage of LUV contents and this effect increased with cell age. Addition of polylysine to the reaction mixture increased even further the leakage of the LUV contents. The cell-free medium of the cell suspension also affected the integrity of the LUV. Cell-free medium, by itself, promoted leakage of LUV contents and caused a reduction in the leakage exerted by polylysine. Centrifugation (8000g) of the cell-free medium decreased its effect, heat treatment (122°C) did not alter its effect and sonication enhanced it. The effects of the cell-free medium are attributed to the presence of cell wall debris of disintegrated cells. 相似文献
6.
Role of roots in regulating the growth rate and cytokinin content in leaves 总被引:1,自引:0,他引:1
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The role of roots in the enhancement of cytokinin content and leaf growth of Phaseolus vulgaris plants after decapitation and partial defoliation was investigated. Partial excision of the roots of plants which were decapitated above the primary leaf node resulted in a reduction of leaf growth and soluble proteins accumulation in the primary leaves. Roots excision was done at time of decapitation and repeated 8 days later. Endogenous cytokinins, known to be involved in enhancing shoot growth, accumulated in the leaves and stems of decapitated and partially defoliated plants. Lower levels of cytokinins were detected in the leaves of decapitated plants with only a partial root system. The level of cytokinins in the roots of decapitated plants was reduced by partial root excision. The growth and accumulation of cytokinins in leaves were, however, not totally suppressed by removing a large proportion of the roots. At the commencement of the experiment the stem had a higher cytokinin content than both the leaves and roots. This suggests that the stem could be an alternative source of cytokinins to the leaves. The cytokinin complement in the leaves of decapitated plants is not identical to that in the roots. It appears that cytokinins supplied by the roots are metabolized in the leaves, or that alternatively certain cytokinins are synthesized in the leaves themselves. 相似文献
7.
The Agrobacterium tumefaciens virC1 gene product binds to overdrive, a T-DNA transfer enhancer. 总被引:10,自引:4,他引:6
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N Toro A Datta O A Carmi C Young R K Prusti E W Nester 《Journal of bacteriology》1989,171(12):6845-6849
In Agrobacterium tumefaciens, a cis-active 24-base-pair sequence adjacent to the right border of the T-DNA, called overdrive, stimulates tumor formation by increasing the level of T-DNA processing. Recent results from our laboratory have suggested that the virC operon which enhances T-DNA processing probably does so because the VirC1 protein interacts with overdrive (N. Toro, A. Datta, M. Yanofsky, and E. W. Nester, Proc. Natl. Acad. Sci. USA 85:8558-8562, 1988). We report here the purification of the VirC1 protein from cells of Escherichia coli harboring a plasmid containing the coding sequences of the virC locus of the octopine Ti plasmid. By gel mobility shift and DNase I footprinting assays, we showed that this purified virC1 gene product binds to overdrive but not to the right border of T-DNA. 相似文献
8.
The influence of specific L- to D-amino acid substitutions on the interaction of pardaxin, a shark repellent neurotoxin polypeptide, with phospholipid vesicles and human erythrocytes is described. Twelve modified, truncated, or fluorescently labeled [with the fluorophore 7-nitrobenz-2-oxa-1,3-diazole-4-yl (NBD) at their N-terminal amino acid] analogues of pardaxin were synthesized by a solid-phase method. Fluorescence measurements were used to monitor the interaction of the analogues with membranes [Rapaport, D., & Shai, Y. (1991) J. Biol. Chem. 266, 23769-23775]. Upon titration of solutions containing the NBD-labeled peptides with small unilamellar vesicles, the fluorescent emission spectra of all NBD-labeled peptides displayed similar blue-shifts, in addition to enhanced intensities, upon relocation of the probe to the more apolar environment. Binding isotherms were constructed from which surface partition constants, in the range of 10(4) M-1, were derived. The existence of an aggregation process, suggested by the shape of the binding isotherms, could be associated only with those analogues in which the N-helix (residues 1-9) was not perturbed. The alpha-helical content of the analogues was estimated by circular dichroism (CD) spectroscopy, both before and after binding to vesicles at neutral pH. The ability of the peptides to dissipate a diffusion potential and to cause calcein release, as well as to lyse human erythrocytes, served to functionally characterize the peptides. The results support a two alpha-helix model, with a bend at position 13, as best describing pardaxin in its membrane-bound state.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
9.
Interaction of antimicrobial dermaseptin and its fluorescently labeled analogues with phospholipid membranes. 总被引:16,自引:0,他引:16
Dermaseptin, a 34 amino-acid residue antimicrobial polypeptide [Mor, A., Nguyen, V. H., Delfour, A., Migliore-Samour, D., & Nicolas, P. (1991) Biochemistry 30, 8824-8830] was synthesized and selectively labeled at its N-terminal amino acid with either 7-nitrobenz-2-oxa-1,3-diazole-4-yl (NBD), rhodamine, or fluorescein. The fluorescent emission spectra of the NBD-labeled dermaseptin displayed a blue-shift upon binding to small unilamellar vesicles (SUV), reflecting the relocation of the fluorescent probe to an environment of increased apolarity. Titrations of solutions containing NBD-labeled dermaseptin with SUV composed of zwitterionic or acidic phospholipids were used to generate binding isotherms, from which were derived surface partition constants of (0.66 +/- 0.06) x 10(4) M-1 and (2.8 +/- 0.3) x 10(4) M-1, respectively. The shape of the binding isotherms, as well as fluorescence energy transfer measurements, suggests that some aggregation of membrane-bound peptide monomers occurs in acidic but not in zwitterionic vesicles. The preferential susceptibility of the peptide to proteolysis when bound to zwitterionic but not to acidic SUV suggests that these aggregates might then penetrate a relatively short distance into the hydrophobic region of the acidic membrane. Furthermore, the results provide good correlation between the peptide's strong binding and its ability to permeate membranes composed of acidic phospholipids, as revealed by a dissipation of diffusion potential and a release of entrapped calcein from SUV. 相似文献
10.
A 24-amino acid peptide corresponding to the S4 segment of the sodium channel was synthesized. In order to perform fluorescence energy transfer measurements and to monitor the interaction of the peptide with lipid vesicles, the peptide was selectively labeled with fluorescence probes at either its N- or C-terminal amino acids. The fluorescent emission spectra of 7-nitrobenz-2-oxa-1,3-diazol-4- yl-(NBD-)labeled analogues displayed blue shifts upon binding to small unilamellar vesicles (SUV), reflecting the relocation of the fluorescent probe to an environment of increased apolarity. The results revealed that both the N- and C-terminus of the S4 segment are located within the lipid bilayer. Titration of solutions containing NBD-labeled peptides with SUV was used to generate binding isotherms, from which surface partition constants, in the range of 10(4) M-1, were derived. The shape of the binding isotherms as well as fluorescence energy transfer measurements suggest that aggregation of peptide monomers within the membrane readily occurs in acidic but not in zwitterionic vesicles. Furthermore, the results provide good correlation between the incidence of aggregation in PC/PS vesicles and the ability of the peptides to permeate the vesicle's membrane. However, a transmembrane diffusion potential had no detectable effect on the location of the peptide within the lipid bilayer or on its aggregation state.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献