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51.
Molecular cloning and functional expression of bacteriophage PK1E-encoded endoneuraminidase Endo NE 总被引:10,自引:3,他引:7
Rita Gerardy-Schahn rea Bethe Thomas Brennecke † Martina Mühlenhoff Matthias Eckhardt Stefan Ziesing Friedrich Lottspeich Matthias Frosch 《Molecular microbiology》1995,16(3):441-450
Homopolymeric α-2,8-linked sialic acid (PSA) has been found as a capsular component of sepsis- and meningitis-causing bacterial pathogens, and on eukaryotic cells as a post-translational modification of the neural cell adhesion molecule (NCAM). The polysaccharide is specifically recognized and degraded by a phage-encoded enzyme, the endo-N-acetylneuraminidase E (Endo NE). Endo NE therefore has become a valuable tool in the study of bacterial pathogenesis and eukaryotic morphogenesis. In this report we describe the molecular cloning of Endo NE and the expression of a functionally active recombinant enzyme. The cloned DNA sequence (2436 bp) encodes a polypeptide of 811 amino acids, which at the 5′ end contains a totally conserved neuraminidase motif. Expressed in Escherichia coli, the enzyme migrates as a single band of approximately 74 kDa in SDS-PAGE. A central domain of 669 amino acid residues is about 90% homologous to the recently cloned Endo NF. Both phage-induced lysis of bacteria and the catalysis of PSA degradation by the recombinant enzyme are efficiently inhibited by a polyclonal antiserum raised against the intact phage particle. The C-terminal region seems to be essential to enzymatic functions, as truncation of 32 amino acids outside the homology domain completely abolishes Endo NE activity. Our data also indicate that the 38 kDa protein, previously assumed to be a subunit of the Endo NE holoenzyme, is the product of a separate gene locus and is not necessary for in vitro depolymerase activity. 相似文献
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John A. Wiens 《Oecologia》1977,31(2):135-151
Summary The energy demands and general food consumption rates of bird populations breeding in North American grasslands are estimated using a simulation model which employs information on population natural history and individual metabolism gathered from several study locations. The total breeding season energy demand of the grassland/shrub-steppe avifaunas ranged from 0.89 kcal m-2 season-1 in arid shrub-steppe to 2.92 kcal m-2 season-1 in a mesic tallgrass prairie. There was substantial variation between years and between census plots, however, and in general the average avian community energy demands did not differ significantly over the range of locations. Production accounted for 0.9 to 1.5% of the total seasonal energy demand. Roughly 11 to 18% of the seasonal energy flow was required in the production of eggs and maintenance and growth of nestlings and fledglings.On the average, between 209 and 386 kg dry wt km-2 of prey were consumed by the bird communities breeding in the grassland locations. Seeds contributed more to the total biomass consumed at the drier plots, but in general, animal prey types comprised roughly 80% of the total biomass eaten. Phytophagous insects were the major component of the animal prey.These low magnitudes of energy flow and biomass consumption attest to the relatively minor role of birds in the processing of energy and biomass in grassland ecosystems. If these populations do play an importnat role in the functioning of grassland ecosystems, it must be quite subtle and indirect. 相似文献
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Several strains of mice bearing the H-2Kk allele were found to generate in vivo strong CTL responses against TNP-haptenated syngeneic cells, while several other strains of mice were found to generate comparably weak or no responses. C3H × DBA/2)F1 mice (H-2k × H-2d) and A/J mice with the recombinant haplotype generated CTL responses in vivo that were completely restricted toward the H-2k haplotype or the K end of the haplotype, respectively. The CTL activity of C3H × DBA/2)F1 and A/J mice against haptenated H-2k targets was found to be more than 25-fold higher than the CTL activity on H-2d targets. The CTL responses in vitro under macroculture conditions showed, on the other hand, only a 3- to 6-fold higher cytotoxic activity against the haptenated H-2k targets as compared with haptenated allogeneic or H-2d targets; and limiting dilution experiments in microcultures revealed that the CTL precursor frequencies were only 2- to 3-fold smaller for TNP-haptenated H-2d or haptenated allogeneic targets than for haptenated H-2k target cells. This indicated that sufficient numbers of H-2d-restricted and allorestricted CTL precursors were actually present in these strains, but did not develop detectable cytotoxic activity in vivo. The exceptional property of the H-2k haplotype is, therefore, only partly determined by a difference in the CTL precursor frequencies, and to the larger extent determined at the level of the activation of the CTL response. 相似文献
55.
Electrical and structural properties of crayfish claw motoneurons in an isolated claw-ganglion preparation 总被引:2,自引:0,他引:2
Theodore J. Wiens 《Journal of comparative physiology. A, Neuroethology, sensory, neural, and behavioral physiology》1976,112(2):213-233
1. | An isolated claw-ganglion preparation of the crayfish is described in which reflex activity is maintained for eight hours or more. |
2. | Intracellular recording and cobalt injection have made it possible to locate and characterize the claw motoneurons. Soma recordings show attenuated axon spikes of 5–10 mV amplitude and subthreshold potentials of the same magnitude. |
3. | The fast closer excitor (FCE) receives subthreshold excitation in parallel with the slow closer excitor (SCE) and the opener inhibitor (OI) even though it seldom fires. |
4. | IPSP's are recorded in the opener excitor (OE) concurrent with OI spikes. |
5. | Cobalt injection reveals a parallel dendritic structure among the three synergists FCE, SCE and OI, and a distinctly different dendritic distribution for OE. The soma siza ranking: OI<>FCE = SCE is apparent. |
6. | The results are discussed and compared with other arthropod systems with regard to the relation between soma size and electrical functions, distribution of somata and dendrites, and effects of cobalt on electrical functioning. A conflict with previous work on this system is discussed. |
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Human mesenchymal stem cells in contact with their environment: surface characteristics and the integrin system 总被引:8,自引:0,他引:8
Docheva D Popov C Mutschler W Schieker M 《Journal of cellular and molecular medicine》2007,11(1):21-38
The identification of mesenchymal stem cells (MSCs) in adult human tissues and the disclosure of their self-renew-al and multi-lineage differentiation capabilities have provided exciting prospects for cell-based regeneration and tis-sue engineering. Although a considerable amount of data is available describing MSCs, there is still lack of information regarding the molecular mechanisms that govern their adhesion and migration. In this work, we will review the current state of knowledge on integrins and other adhesion molecules found to be expressed on MSCs. The discussed topics include the characteristics of MSCs and their clinical applications, integrins and their central role in cell-matrix attachment and migration, and comments on mobilization, differentiation and contribution to tumour development. Finally, by understanding the complex and fundamental pathways by which MSCs attach and migrate, it might be possible to fine-tune the strategies for effective and safe use of MSCs in regenerative therapies. 相似文献
60.
Almost all large-scale projects in mass spectrometry-based proteomics use trypsin to convert protein mixtures into more readily analyzable peptide populations. When searching peptide fragmentation spectra against sequence databases, potentially matching peptide sequences can be required to conform to tryptic specificity, namely, cleavage exclusively C-terminal to arginine or lysine. In many published reports, however, significant numbers of proteins are identified by non-tryptic peptides. Here we use the sub-parts per million mass accuracy of a new ion trap Fourier transform mass spectrometer to achieve more than a 100-fold increased confidence in peptide identification compared with typical ion trap experiments and show that trypsin cleaves solely C-terminal to arginine and lysine. We find that non-tryptic peptides occur only as the C-terminal peptides of proteins and as breakup products of fully tryptic peptides N-terminal to an internal proline. Simulating lower mass accuracy led to a large number of proteins erroneously identified with non-tryptic peptide hits. Our results indicate that such peptide hits in previous studies should be re-examined and that peptide identification should be based on strict trypsin specificity. 相似文献