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51.
Localization of a series of RNA-protein cross-link sites in the 23S and 5S ribosomal RNA from Escherichia coli, induced by treatment of 50S subunits with three different bifunctional reagents. 总被引:13,自引:4,他引:9
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50S ribosomal subunits were reacted with bis-(2-chloroethyl)methylamine, 2-iminothiolane or methyl p-azidophenyl acetimidate, and RNA-protein cross-link sites on the RNA were localised using our published procedures. The degree of precision with which these sites could be determined was variable, depending on the particular protein or RNA region concerned. The following positions in the 23S RNA were identified as encompassing the individual cross-link sites (numbered from the 5'-end, with asterisks denoting sites previously reported): L1, 1864-67, 1876-78, 2119-33, 2163-72*, L2, 1819-20*; L3, 2832-34; L4, 320-25*; 613-17*; L5, 2307; L6, 2473-81*; L9, 1484-91; L11, 1060-62; L13, 547-50; L14, 1993-2002; L17, 1260-95; L18, 2307-20; L19, 1741-58; L21, 544-48*; 1198-1248; L23, 63-65, 137-41*; L24, 99-107*; L27, 2272-83, 2320-23*; 2332-37*; L28, 195-242, 368-424; L29, 101-02*; L30, 931-38; L32, 2878-90; L33, 2422-24. Cross-links to 5S RNA were observed with L5 (positions 34-41), and L18 (precise site not localised). 相似文献
52.
Localisation of a series of intra-RNA cross-links in the secondary and tertiary structure of 23S RNA, induced by ultraviolet irradiation of Escherichia coli 50S ribosomal subunits. 总被引:19,自引:9,他引:10
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Intra-RNA cross-links were introduced into E. coli 50S ribosomal subunits by mild ultraviolet irradiation. The subunits were partially digested with cobra venom nuclease, and the cross-linked RNA complexes were isolated by two-dimensional electrophoresis. Many of the complexes were submitted to a second partial digestion procedure. Oligonucleotide analysis of the RNA fragments obtained in this manner enabled cross-links between the following ribonuclease T1 oligonucleotides in the 23S RNA to be established: positions 292-296 and 339-350; 601-604 and 652-656; 1018-1022 and 1140-1149; 1433-1435 and 1556-1560; 1836-1839 and 1898-1903; 2832-2834 (tentative) and 2878-2885; 2849-2852 and 2865-2867 (tentative); 739-748 and 2609-2618; 571-577 and 2030-2032; 1777-1792 (tentative) and 2584-2588. The first seven of these cross-links lie within the secondary structure of the 23S RNA, whereas the last three are tertiary structural cross-links. The degree of precision of the individual determinations was variable, depending on the nucleotide sequence in the vicinity of the cross-link site concerned. 相似文献
53.
A detailed study of courtship in Spodoptera littoralis showed that there were four significant behaviour patterns. The male flew to a calling female and hovered above her with his brushes fully extended. In response, the female lifted her wings, curved her abdomen and withdrew her pheromone gland. The male settled beside the female to pair and then moved to hang head downwards during copulation. Thirty percent of successful courtships lacked one of the main behaviour patterns. Nearly half the courtships observed did not end in copulation: none of these included all of four major behaviour patterns and the majority lacked two or three. Females often rejected males with a rapid flick of the wings. Antennaless males did not mate or extend the brushes in response to a calling female. Just over half of the antennaless females observed during 135-min tests mated with normal males, but courtship was abnormal. Olfactory cues appeared to be important to females in recognizing the courting male, since antennaless females did not wing flick, were significantly more likely to take to flight as the result of the male brush display and frequently failed to retract the pheromone gland during the latter states of courtship. The courtship behaviour of S. littoralis is compared with published accounts for other Noctuids. 相似文献
54.
Heterogeneous binding of high mobility group chromosomal proteins to nuclei 总被引:7,自引:5,他引:2
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A dramatic difference is observed in the intracellular distribution of the high mobility group (HMG) proteins when chicken embryo fibroblasts are fractionated into nucleus and cytoplasm by either mass enucleation of cytochalasin-B-treated cells or by differential centrifugation of mechanically disrupted cells. Nuclei (karyoplasts) obtained by cytochalasin B treatment of cells contain more than 90 percent of the HMG 1, while enucleated cytoplasts contain the remainder. A similar distribution between karyoplasts and cytoplasts is observed for the H1 histones and the nucleosomal core histones as anticipated. The presence of these proteins, in low amounts, in the cytoplast preparation can be accounted for by the small percentage of unenucleated cells present. In contrast, the nuclei isolated from mechanically disrupted cells contain only 30-40 percent of the total HMGs 1 and 2, the remainder being recovered in the cytosol fraction. No histone is observed in the cytosol fraction. Unike the higher molecular weight HMGs, most of the HMGs 14 and 17 sediment with the nuclei after cell lysis by mechanical disruption. The distribution of HMGs is unaffected by incubating cells with cytochalasin B and mechanically fractionating rather than enucleating them. Therefore, the dramatic difference in HMG 1 distribution observed using the two fractionation techniques cannot be explained by a cytochalasin-B-induced redistribution. On reextraction and sedimentation of isolated nuclei obtained by mechanical cell disruption, only 8 percent of the HMG 1 is released to the supernate. Thus, the majority of the HMG 1 originally isolated with these nuclei, representing 35 percent of the total HMG 1, is stably bound, as is all the HMGs 14 and 17. The remaining 65 percent of the HMGs 1 and 2 is unstably bound and leaks to the cytosol fraction under the conditions of mechanical disruption. It is suggested that the unstably bound HMGs form a protein pool capable of equilibrating between cytoplasm and stably bound HMGs. 相似文献
55.
56.
57.
RNA-protein cross-linking in Escherichia coli 30S ribosomal subunits; determination of sites on 16S RNA that are cross-linked to proteins S3, S4, S5, S7, S8, S9, S11, S13, S19 and S21 by treatment with methyl p-azidophenyl acetimidate. 总被引:7,自引:4,他引:3
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M Osswald B Greuer R Brimacombe G St?ffler H B?umert H Fasold 《Nucleic acids research》1987,15(8):3221-3240
RNA-protein cross-links were introduced into E. coli 30S ribosomal subunits by treatment with methyl p-azidophenyl acetimidate. After partial nuclease digestion of the RNA moiety, a number of cross-linked RNA-protein complexes were isolated by a new three-step procedure. Protein and RNA analysis of the individual complexes gave the following results: Proteins S3, S4, S5 and S8 are cross-linked to the 5'-terminal tetranucleotide of 16S RNA. S5 is also cross-linked to the 16S RNA within an oligonucleotide encompassing positions 559-561. Proteins S11, S9, S19 and S7 are cross-linked to 16S RNA within oligonucleotides encompassing positions 702-705, 1130-1131, 1223-1231 and 1238-1240, respectively. Protein S13 is cross-linked to an oligonucleotide encompassing positions 1337-1338, and is also involved in an anomalous cross-link within positions 189-191. Protein S21 is cross-linked to the 3'-terminal dodecanucleotide of the 16S RNA. 相似文献
58.
Covalent cross-linking of poly(A) to Escherichia coli ribosomes, and localization of the cross-link site within the 16S RNA. 总被引:7,自引:3,他引:4
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Poly(A) can be cross-linked to E. coli 70S ribosomes in the presence of tRNALys by mild ultraviolet irradiation. The cross-linking reaction is exclusively with the 30S subunit, and involves primarily the RNA moiety. Following a partial nuclease digestion, cross-linked complexes containing poly(A) and fragments of the 16S RNA were isolated by affinity chromatography on oligo(dT)-cellulose. The complexes were purified by gel electrophoresis and subjected to oligonucleotide analysis, which revealed a single cross-link site within positions 1394-1399 of the 16S RNA. The same pattern of cross-linking, at about one-fifth of the intensity, was observed in the absence of tRNALys. The cross-link site to poly(A), together with other sites in the 16S RNA that have been implicated in ribosomal function, is discussed in the framework of our recent model for the three-dimensional structure of 16S RNA; all of the functional sites are clustered together in two distinct groups in the model. 相似文献
59.
A model for the spatial arrangement of the proteins in the large subunit of the Escherichia coli ribosome. 总被引:12,自引:2,他引:10
J Walleczek D Schüler M Stffler-Meilicke R Brimacombe G Stffler 《The EMBO journal》1988,7(11):3571-3576
A three-dimensional model for the arrangement of 29 of the 33 proteins from the Escherichia coli large ribosomal subunit has been generated by interactive computer graphics. The topographical information that served as input in the model building process was obtained by combining the immunoelectron microscopically determined network of epitope-epitope distances on the surface of the large ribosomal subunit with in situ protein-protein cross-linking data. These two independent sets of data were shown to be compatible by geometric analysis, thus allowing the construction of an inherently consistent model. The model shows (i) that the lower third of the large subunit is protein-poor, (ii) that proteins known to be functionally involved in peptide bond formation and translocation are clustered in two separate regions, (iii) that proteins functionally interdependent during the self-assembly of the large subunit are close neighbours in the mature subunit and (iv) that proteins forming the early assembly nucleus are grouped together in a distinct region at the 'back' of the subunit. 相似文献
60.