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Demonstration that a chemically synthesized BPV1 oncoprotein and its C-terminal domain function to induce cellular DNA synthesis 总被引:17,自引:0,他引:17
Bovine papillomavirus type 1 contains the smallest known oncogene (ORF E5), encoding a hydrophobic 44 amino acid protein. To study the biochemical functions of the E5 oncoprotein, we have chemically synthesized it and several deletion mutant peptides. We demonstrate induction of cellular DNA synthesis in growth-arrested cells by microinjection of E5 oncoprotein. This activity can be broken down into two functionally distinguishable domains. Remarkably, the first domain, which alone is sufficient to induce cellular DNA synthesis, contains only the C-terminal 13 amino acids. This is the smallest known protein fragment that can autonomously activate cellular DNA synthesis. The second domain is the hydrophobic middle region, which by itself fails to induce cellular DNA synthesis but confers a 1000-fold increase in specific activity. The N-terminal one-third of the molecule is dispensable for induction of DNA synthesis. 相似文献
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Summary The results of a recent quantitative analysis of the Teorell membrane oscillator are utilized to explore its role as an excitability analogue. Special attention is paid to its role as a mechano-electric transducer. A membrane of exceptionally well-defined pore structure has been used in this study. The analogue properties arise from nonlinear coupling between water and salt fluxes. When the membrane is simultaneously subjected to controlled gradients of hydrostatic pressure, electrical potential and concentration, bi-stable stationary states can be produced. These arise from the opposing effects of pressure and electro-osmosis on the volume flow. Transitions between these states show hysteresis. The factors governing such transitions are analogous to certain types of stimuli encountered in the natural excitation process. The membrane system also shows oscillatory behavior when the hydrostatic pressure gradient is allowed to vary under constant current conditions. This property is related to the bi-stable stationary state phenomena and is compared to the regenerative behavior found in biologically excitable tissues. Particular emphasis is placed upon analogies between the membrane oscillator and certain natural tissues. The importance of the nonlinear nature of the force-flux coupling in the analogue is stressed, and its possible relevance to biological excitability indicated. Some consideration is also given to the role of electro-osmotic flux coupling in biological tissues. 相似文献
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(Z)- and (E)-4-amino-2-(trifluoromethyl)-2-butenoic acid (4 and 5, respectively) were synthesized and investigated as potential mechanism-based inactivators of gamma-aminobutyric acid aminotransferase (GABA-AT) in a continuing effort to map the active site of this enzyme. The core alpha-trifluoromethyl-alpha,beta-unsaturated ester moiety was prepared via a Reformatsky/reductive elimination coupling of the key intermediates tert-butyl 2,2-dichloro-3,3,3-trifluoropropionate and N,N-bis(tert-butoxy-carbonyl)glycinal. Both 4 and 5 inhibited GABA-AT in a time-dependent manner, but displayed non-pseudo-first-order inactivation kinetics; initially, the inactivation rate increased with time. Further investigation demonstrated that the actual inactivator is generated enzymatically from 4 or 5. This inactivating species is released from the active site prior to inactivation, and as a result, 4 and 5 cannot be defined as mechanism-based inactivators. Furthermore, 4 and 5 are alternate substrates for GABA-AT, transaminated by the enzyme with Km values of 0.74 and 20.5 mM, respectively. Transamination occurs approximately 276 and 305 times per inactivation event for 4 and 5, respectively. The enzyme also catalyzes the elimination of the fluoride ion from 4 and 5. A mechanism to account for these observations is proposed. 相似文献
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