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1.
2.
Maintenance of the cellobiose utilization genes of Escherichia coli in a cryptic state 总被引:6,自引:1,他引:5
The genes for cellobiose utilization are normally cryptic in Escherichia
coli. The cellobiose system was used as a model to understand the process
by which silent genes are maintained in microbial populations. Previously
reported was (1) the isolation of a mutant strain that expresses the
cellobiose-utilization (Cel) genes and (2) that expression of those genes
allows utilization of three beta- glucoside sugars: cellobiose, arbutin,
and salicin. The Cel gene cluster has now been cloned from that mutant
strain. In the course of locating the Cel genes within the cloned DNA
segment, it was discovered that inactivation of the Cel-encoded hydrolase
rendered the host strain sensitive to all three beta-glucosides as potent
inhibitors. This sensitivity arises from the accumulation of the
phosphorylated beta- glucosides. Because even the fully active genes
conferred some degree of beta-glucoside sensitivity, the effects of
cellobiose on a series of five Cel+ mutants of independent origin were
investigated. Although each of those strains utilizes cellobiose as a sole
carbon and energy source, cellobiose also acts as a potent inhibitor that
reduces the growth rate on glycerol 2.5-16.5-fold. On the other hand,
wild-type strains that cannot utilize cellobiose are not inhibited. The
observation that the same compound can serve either as a nutrient or as an
inhibitor suggests that, under most conditions in which cellobiose will be
present together with other resources, there is a strong selective
advantage to having the cryptic (Cel0) allele. In those environments in
which cellobiose is the sole, or the best, resource, mutants that express
the genes (Cel+) will have a strong selective advantage. It is suggested
that temporal alternation between these two conditions is a major factor in
the maintenance of these genes in E. coli populations. This alternation of
environments and fitnesses was predicted by the model for cryptic-gene
maintenance that was previously published.
相似文献
3.
The inhibition of pepsin-catalysed reactions by products and product analogues. Kinetic evidence for ordered release of products 总被引:4,自引:4,他引:0 下载免费PDF全文
1. The inhibition of pepsin-catalysed hydrolysis of N-acetyl-l-phenylalanyl-l-phenylalanylglycine by products and product analogues was studied. 2. The non-competitive nature of the inhibition by the product N-acetyl-l-phenylalanine confirms an ordered release of products, and points to a common mechanism (involving an amino-enzyme) for pepsin-catalysed transpeptidation and hydrolysis reactions. 3. N-Acetyl-l-phenylalanine ethyl ester is also a non-competitive inhibitor, but here the inhibition is of the ;dead-end' type. No ethanol is detectable in reaction mixtures, indicating that this ester cannot act as an amino group acceptor in a transpeptidation process. 4. The same is true for N-methanesulphonyl-l-phenylalanine methyl and methyl thiol esters. No methanethiol is liberated when the methyl thiol ester is present as an inhibitor of the hydrolytic reaction, and the hope that such a thiol ester would effectively trap the amino-enzyme was not fulfilled. 相似文献
4.
Biophysical characterization of recombinant proteins expressing the leucine zipper-like domain of the human immunodeficiency virus type 1 transmembrane protein gp41. 总被引:2,自引:2,他引:0 下载免费PDF全文
Envelope oligomerization is thought to serve several crucial functions during the life cycle of human immunodeficiency virus type 1 (HIV-1). We recently reported that virus entry requires coiled-coil formation of the leucine zipper-like domain of the HIV-1 transmembrane envelope glycoprotein gp41 (C. Wild, T. Oas, C. McDanal, D. Bolognesi, and T. Matthews, Proc. Natl. Acad. Sci. USA 89:10537-10541, 1992; C. Wild, J. W. Dubay, T. Greenwell, T. Baird, Jr., T. G. Oas, C. McDanal, E. Hunter, and T. Matthews, Proc. Natl. Acad. Sci. USA 91:12676-12680, 1994). To determine the oligomeric state mediated by this region of the envelope, we have expressed the zipper motif as a fusion partner with the monomeric maltose-binding protein of Escherichia coli. The biophysical properties of this protein were characterized by velocity and equilibrium sedimentation, size exclusion chromatography, light scattering, and chemical cross-linking analyses. Results indicate that the leucine zipper sequence from HIV-1 is capable of multimerizing much larger and otherwise monomeric proteins into extremely stable tetramers. Recombinant proteins containing an alanine or a serine substitution at a critical isoleucine residue within the zipper region were also generated and similarly analyzed. The alanine- and serine-substituted proteins behaved as tetrameric and monomeric species, respectively, consistent with the influence of these same substitutions on the helical coiled-coil structure of synthetic peptide models. On the basis of these findings, we propose that the fusogenic gp4l structure involves tetramerization of the leucine zipper domain which is situated approximately 30 residues from the N-terminal fusion peptide sequence. 相似文献
5.
Zhan Wang Suzon Larocque Evgeny Vinogradov Jean-Robert Brisson Andrew Dacanay Marshall Greenwell Laura L Brown Jianjun Li Eleonora Altman 《European journal of biochemistry》2004,271(22):4507-4516
Aeromonas salmonicida is a pathogenic aquatic bacterium and the causal agent of furunculosis in salmon. In the course of this study, it was found that when grown in vitro on tryptic soy agar, A. salmonicida strain 80204-1 produced a capsular polysaccharide with the identical structure to that of the lipopolysaccharide O-chain polysaccharide. A combination of 1D and 2D NMR methods, including a series of 1D analogues of 3D experiments, together with capillary electrophoresis-electrospray MS (CE-ES-MS), compositional and methylation analyses and specific modifications was used to determine the structure of these polysaccharides. Both polymers were shown to be composed of linear trisaccharide repeating units consisting of 2-acetamido-2-deoxy-D-galacturonic acid (GalNAcA), 3-[(N-acetyl-L-alanyl)amido]-3,6-dideoxy-D-glucose[3-[(N-acetyl-L-alanyl)amido]-3-deoxy-D-quinovose, Qui3NAlaNAc] and 2-acetamido-2,6-dideoxy-D-glucose (2-acetamido-2-deoxy-D-quinovose, QuiNAc) and having the following structure: [-->3)-alpha-D-GalpNAcA-(1-->3)-beta-D-QuipNAc-(1-->4)-beta-D-Quip3NAlaNAc-(1-]n, where GalNAcA is partly presented as an amide and AlaNAc represents N-acetyl-L-alanyl group. CE-ES-MS analysis of CPS and O-chain polysaccharide confirmed that 40% of GalNAcA was present in the amide form. Direct CE-ES-MS/MS analysis of in vivo cultured cells confirmed the formation of a novel polysaccharide, a structure also formed in vitro, which was previously undetectable in bacterial cells grown within implants in fish, and in which GalNAcA was fully amidated. 相似文献
6.
Stabilization of Microsatellite Sequences by Variant Repeats in the Yeast Saccharomyces Cerevisiae 总被引:2,自引:0,他引:2 下载免费PDF全文
We examined the effect of a single variant repeat on the stability of a 51-base pair (bp) microsatellite (poly GT). We found that the insertion stabilizes the microsatellite about fivefold in wild-type strains. The stabilizing effect of the variant base was also observed in strains with mutations in the DNA mismatch repair genes pms1, msh2 and msh3, indicating that this effect does not require a functional DNA mismatch repair system. Most of the microsatellite alterations in the pms1, msh2 and msh3 strains were additions or deletions of single GT repeats, but about half of the alterations in the wild-type and msh6 strains were large (>8 bp) deletions or additions. 相似文献
7.
Protein synthesis by hybrid ribosomes reconstructed from rabbit reticulocyte ribosomal core-particles and amphibian or fungal split-proteins. 总被引:1,自引:0,他引:1 下载免费PDF全文
It was shown that high-salt (2.75 M-NH4Cl/69mM-MgCl2) shock treatment at 0 degrees C of the larger subparticles (L-subparticles) of rabbit, Xenopus laevis and Neurospora crassa cytoplasmic ribosomes yielded split-protein fractions that were not only functionally equivalent but also interchangeable. Thus, although the remaining core-particles were inactive in both the puromycin reaction and in poly(U)-directed polyphenylalanine synthesis, activity was restored when they were combined with either homologous or heterologous split-protein fractions. This technique was used to prepare active hybrid L-subparticles, e.g. rabbit cores/N. crassa split-proteins, and also active hybrid ribosomes, e.g. rabbit smaller subparticle/X. laevis core-particle/rabbit split-proteins. Rabbit and X. laevis split-protein fractions labelled with 14C by reductive methylation with [14C]formaldehyde and sodium cyanoborohydride were both shown to bind to rabbit core-particles in approximate correlation with the degree of re-activation. The split-protein fractions of rabbit and X. laevis L-subparticles were analysed by two-dimensional and sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. The molecular weights (measured in sodium dodecyl sulphate gels) of the split-proteins of rabbit and X. laevis L-subparticles were found to be similar. These results demonstrate that the peptidyltransferase active centre of cytoplasmic ribosomes of eukaryotes has components that are interchangeable over a wide evolutionary range. Evidently the essential molecular architecture of the active centre is highly conserved. 相似文献
8.
9.
Hengshan Zhang Ane F. B. Zeidler Wei Song Christopher M. Puccia Ewa Malc Patricia W. Greenwell Piotr A. Mieczkowski Thomas D. Petes Juan Lucas Argueso 《Genetics》2013,193(3):785-801
The kinetochore is the macromolecular protein complex that mediates chromosome segregation. The Dsn1 component is crucial for kinetochore assembly and is phosphorylated by the Aurora B kinase. We found that Aurora B phosphorylation of Dsn1 promotes the interaction between outer and inner kinetochore proteins in budding yeast. 相似文献