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101.
Amber mutations in Escherichia coli essential genes: isolation of mutants affected in the ribosomes.
Geneviève Delcuve Teresa Cabezón Alain Ghysen Albert Herzog Alex Bollen 《Molecular & general genetics : MGG》1977,157(2):149-153
Summary A method to obtain amber mutations in ribosomal protein genes is described. It relies on the P1-mediated localized mutagenesis (Hong and Ames, 1971) and on the fact that the recipient strain contains (a) an efficient but genetically unstable suppressor, (b) a particular thermoinducible prophage which kills suppressor hosts at 42° C. Exposure of these bacteria to the high temperature yields frequent suppressor-free derivatives while none will be found if the strain carries an amber mutation in an essential gene. Eleven mutants have been isolated by this method, of which at least six appear to carry amber mutations. All of them map close to, and to the right of spcA, in a region which codes mostly for ribosomal proteins. Three mutants were studied biochemically; all three show defective ribosomal assembly in vivo upon loss of suppression. 相似文献
102.
Teresa Rondon Rota 《In vitro cellular & developmental biology. Plant》1977,13(5):280-292
Summary Trachoma organisms of serotype B were grown serially in irradiated cells (McCoy, BHK-21, Microbiological Associates, and BHK-21,
Lister) and tested for infectivity in monolayers of five mammalian cell lines (BHK-21, CHO, HeLa S3, McCoy and OWMK) and two diploid strains (ST/BTL and WI-38). All cell types had low susceptibility to chlamydial infection
but the number of inclusions increased when the inoculum was centrifuged onto the monolayers, or when the cells were irradiated.
Infection was higher in non-irradiated CHO than in irradiated CHO in three out of a total of six experiments. Inclusion number
was increased 300 times in HeLa S3 and up to three times in the other cell types after treatment with diethylaminoethyl-dextran (DEAE-D). Serial passage of
Chlamydia trachomatis serotype B (strain Har-36) in CO60 McCoy and CO60 BHK-21 Lister resulted in partial adaptation of the strain to the host cell. The phenomenon of adaptation of serotype B to
McCoy compensated for the lower susceptibility of this cell revealed when McCoy cells were inoculated with trachoma elementary
bodies grown in BHK-21 Lister or in chick embryo yolk sac. Trachoma organisms of immunotypes A, B and C prepared in yolk sac
produced more inclusion-forming units per ml in CO60 BHK-21 Lister than in CO60 McCoy.
This research was supported by a grant from the National Eye Institute (EI-00812-08), and by the Arabian American Oil Company.
The paper is dedicated to the memory of Francis B. Gordon, who pioneered research methods for the cultivation of trachoma
and inclusion conjunctivitis (TRIC) agents in cell culture. Dr. Gordon patiently studied tables and photographs which accompany
this text when he visited our laboratory on the day prior to his sailing to England on the ill-fated voyage in which he and
Mrs. Gordon perished (October 1973). 相似文献
103.
The ouabain-insensitive, Mg2+-dependent, Na+-stimulated ATPase activity present in fresh basolateral plasma membranes from guinea-pig kidney cortex cells (prepared at pH 7.2) can be increased by the addition of micromolar concentrations of Ca2+ to the assay medium. The Ca2+ involved in this effect seems to be associated with the membranes in two different ways: as a labile component, which can be quickly and easily ‘deactivated’ by reducing the free Ca2+ concentration of the assay medium to values lower than 1 μM; and as a stable component, which can be ‘deactivated’ by preincubating the membranes for periods of 3–4 h with 2 mM EDTA or EGTA. Both components are easily activated by micromolar concentrations of Ca2+. The of the system for Na+ is the same, 8 mM, whether only the stable component or both components, stable and labile, are working. In other words, the activating effect of Ca2+ on the Na+-stimulated ATPase is on the , and not on the of the system for Na+. The activating effect of Ca2+ may be related to some conformational change produced by the interaction of this ion with the membranes, since it can also be obtained by resuspending the membranes at pH 7.8 or by ageing the preparations. Changes in the Ca2+ concentration may modulate the ouabain-insensitive, Na+-stimulated ATPase activity. This modulation could regulate the magnitude of the extrusion of Na+ accompanied by Cl? and water that these cells show, and to which the Na+-ATPase has been associated as being responsible for the energy supply of this mode of Na+ extrusion. 相似文献
104.
Alexander A. Kortt John E. Burns J. Bruce Caldwell Teresa Ferro Phillip M. Strike 《The protein journal》1991,10(2):183-188
The primary structure of acidic trypsin inhibitor-2a (WBTI-2a,pI 5.9) fromPsophocarpus tetragonolobus (L.) DC seed was determined. This inhibitor consists of a single polypeptide chain of 180 amino acids including four half-cystine residues and has an N-terminal residue of pyroglutamic acid. The sequence of WBTI-2a,pI 5.9, showed 84% identity to acidic trypsin inhibitor-2 (WBTI-2,pI 5.1) but only 57% identity to the basic trypsin inhibitor (WBTI-1,pI 8.9) and 50% identity to the chymotrypsin inhibitor of winged bean. The data indicate that winged bean seed contains a family of three Kunitz-type inhibitors which have about 50% identity. 相似文献
105.
Immaculada Herrero María Teresa Miras-Portugal José Sánchez-Prieto 《Journal of neurochemistry》1992,59(4):1574-1577
Abstract: The effects of arachidonic acid and phorbol esters in the Ca2+ -dependent release of glutamate evoked by 4-aminopyridine (4-AP) in rat cerebrocortical synaptosomes were studied. In the absence of arachidonic acid, high concentrations (500 n M ) of 4β-phorbol dibutyrate (4β-PDBu) were required to enhance the release of glutamate. However, in the presence of arachidonic acid, low concentrations of 4β-PDBu (1–50 n M ) were effective in potentiating glutamate exocytosis. This potentiation of glutamate release by phorbol esters was not observed with the methyl ester of arachidonic acid, which does not activate protein kinase C. Moreover, pretreatment of synaptosomes with the protein kinase inhibitor staurosporine also prevented the stimulatory effect by arachidonic acid and phorbol esters. These results suggest that the activation of protein kinase C by both arachidonic acid and phorbol esters may play a role in the potentiation of glutamate exocytosis. 相似文献
106.
DNA polymerase α/primase (Polα) is the key replication enzyme in eukaryotic cells. This enzyme synthesizes and elongates short
RNA primers at an unwound origin of replication. Polα was used as an affinity ligand to identify cellular replication factors
interacting with it. Protein complexes between Polα and cellular factors were analyzed by co-immunoprecipitations with monoclonal
antibodies directed against Polα and by protein affinity chromatography of cell extracts derived from pure G1-and S-phase cell populations on Polα affinity columns. Co-immunoprecipitations resulted in the identification of a polypeptide
with a molecular weight of 46 kDa. For Polα affinity chromatography, the ligand was purified from insect cells infected with
a recombinant baculovirus encoding the catalytic subunit (p180) of Polα (Copeland and Wang, 1991). With 5×108 infected Sf9 cells, a rapid one step purification protocol was used which yielded in five hours 0.6 mg pure enzyme with a
specific activity of 140,000 units/mg. The G1-and S-phase cell populations were generated by block, release and counterflow centrifugal elutriation of exponentially growing
human MANCA cells. Starting with 2×109 non synchronous cells, 5×108 G1-phase cells were isolated. Chromatography of cell extracts derived from G1-or S-phase cells on Polα affinity columns resulted in identifying several polypeptides in the range of 40–70 kDa. Some of
these polypeptides are more abundant in eluates derived from S-phase extracts than from G1-phase extracts. 相似文献
107.
S. M. Teresa Hernández-Sotomayor Graham Carpenter 《The Journal of membrane biology》1992,128(2):81-89
While EGF has an important function in cell growth regulation, the molecular mechanisms by which intracellular signal connect the EGF: receptor complex on the plasma membrane with the initiation of DNA synthesis and mitogenesis is not well understood. The discovery that rasGAP, PI-3 kinase and PLC-gamma 1 are substrates for the EGF receptor tyrosine kinase has provided a beginning in understanding the biochemistry underlying growth factor receptor transduction. 相似文献
108.
Teresa J. Arrowsmith Francisco Malpartida David H. Sherman Ashley Birch David A. Hopwood John A. Robinson 《Molecular & general genetics : MGG》1992,234(2):254-264
Summary Cloned DNA encoding polyketide synthase (PKS) genes from one Streptomyces species was previously shown to serve as a useful hybridisation probe for the isolation of other PKS gene clusters from the same or different species. In this work, the actI and actIII genes, encoding components of the actinorhodin PKS of Streptomyces coelicolor, were used to identify and clone a region of homologous DNA from the monensin-producing organism S. cinnamonensis. A 4799 by fragment containing the S. cinnamonensis act-homologous DNA was sequenced. Five open reading frames (ORFs 1–5) were identified on one strand of this DNA. The five ORFs show high sequence similarities to ORFs that were previously identified in the granaticin, actinorhodin, tetracenomycin and whiE PKS gene clusters. This allowed the assignment of the following putative functions to these five ORFS : a heterodimeric -ketoacyl synthase (ORF1 and ORF2), an acyl carrier protein (ORF3), a -ketoacyl reductase (ORF5), and a bifunctional cyclase/dehydrase (ORF4). The ORFs are encoded in the order ORFl-ORF2-ORF3-ORF5-ORF4, and ORFs-1 and -2 show evidence for translational coupling. This act-homologous region therefore appears to encode a PKS gene cluster. A gene disruption experiment using the vector pGM 160, and other evidence, suggests that this cluster is not essential for monensin biosynthesis but rather is involved in the biosynthesis of a cryptic aromatic polyketide in S. cinnamonensis. An efficient plasmid transformation system for S. cinnamonensis has been established, using the multicopy plasmids pWOR120 and pWOR125. 相似文献
109.
Summary
Chamaedorea bartlingiana is a dioecious palm that grows in the cloud forest understories of the Venezuelan Andes. Age and sexual differences in phenology and reproductive patterns were studied in labelled individuals of all age categories. This species has long-lived leaves and low leaf production, both characteristic of understory plants. Growth rates are lower in juveniles than in adults and in females than in males, as in other palms. Male and female individuals show different reproductive patterns. Male inflorescences are always produced at the same rate and the probability of surviving until anthesis is constant. Females produce reproductive buds at the same rate as males, but these buds have a 35% probability of becoming a ripe infrutescence if the plant has infrutescences already growing, and 70% if it does not. This pattern and the slow growth of inflorescences (1 year for males from bud to flowers, 2 years for females from bud to ripe fruits) cause a pluriannual reproductive pattern at the population level. Field germination does not follow this pattern, but shows one annual peak probably related to environmental conditions. 相似文献