Search

Our results demonstrate that the GAL4 two-hybrid system can be useful for studying interactions of the wild-type and mutant forms of Ras proteins with the CDC25 guanine nucleotide exchange factor (CDC25-GEF). In addition, our findings show that a negative result in the GAL4 two-hybrid system does not indicate that the two proteins tested do not interact under all conditions but only that they do not interact under the specific conditions examined. We recommend that the two-hybrid system be employed in combination with other approaches, including molecular genetic analyses and in vitro binding experiments, for the study of Ras and CDC25-GEF interactions.

A broad array of therapeutic options is currently available for the management of patients with AV nodal reentrant tachycardia. While acute termination of tachycardias is readily achieved, either by vagal maneuvers or intravenous medication, the decision to embark on a long-term therapeutic plan to prevent recurrences must be clinically individualized. When a chronic pharmacologic approach is desired, electrophysiologic testing is invaluable for confirming the diagnosis and selecting appropriate medication. However, the growing awareness of potential proarrhythmic effects and the inconvenience and expense of lifelong drug therapy, coupled with other advances in the field, have made nonpharmacologic approaches more attractive. This is especially so for symptomatic younger patients. The definitive cure rates achievable with surgery are now being approached by transcatheter AV nodal modification procedures that ablate AV nodal reentrant tachycardia while preserving anterograde AV nodal conduction. Over the next decade, it is likely that the latter technique will become widely used for the long-term management of symptomatic AV nodal reentrant tachycardia.

The bifunctional tryptophan biosynthetic enzyme N-(5'-phosphoribosyl)anthranilate (PRA) isomerase/indole-3-glycerol phosphate (InGP) synthetase was purified from Escherichia coli cultures incubated under several conditions, some of which result in inactivation of the enzyme (starvation for ammonium or sulfate, chloramphenicol inhibition). Recovery of enzyme activity during purification from cultures incubated under inactivating conditions suggested that activity was restored or that an inhibitor was removed. In these preparations, the enzymatic properties were altered slightly but not in a manner that could account for inactivation. Each preparation exhibited one major protein species (Mr approximately 50,000) when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis but revealed several species during isoelectric focusing in the range of pH 5.5 to 6.0. Comparison of the distribution of species indicated that modification to the more acidic forms had occurred in growing and nongrowing cells but not during stationary phase, glucose starvation, or energy depletion. The results suggested that inactivation and modification had occurred in vivo by an energy-dependent process. Several metabolites tested did not inhibit the purified enzyme. Attempts to detect an inhibitor in crude cell extracts were also not successful.

Approximate spectral representations are developed for the memory kernel which characterizes self-diffusion. These spectral representations are based upon approximate eigenfunctions constructed via the Rayleigh variational principle. A heuristic model is developed first in an effort to provide physical insight into the nature of the approximations employed, and then a number of specific trial functions are examined. These trial functions include sums of identical one- and two-particle functions as well as linear combinations of hydrodynamical variables. The results from these spectral representations indicate that the long-time behavior of the memory kernel (and thereby of the momentum autocorrelation function) is sensitive to the long-range effects of the interparticle potential. In addition, the equivalence of most of these spectral representations to specific low-order perturbation approximations is demonstrated.

The metabolism of 184 individual proteins was examined in exponentially growing Escherichia coli. Cells were labeled with [3H]leucine and [35S]methionine using either a pulse-chase or a continuous labeling method. Proteins were fractionated by two-dimensional gel electrophoresis and their stabilities relative to total protein were determined from the 3H/35S ratios. Forty-seven proteins appeared to be unstable and were probably either degraded or modified to electrophoretically distinct forms. The apparent half-lives of these proteins calculated from the pulse-chase data varied from 2.0 to 23 h. Twenty-seven proteins appeared to be products of post-translational modifications. One hundred fourteen proteins appeared to be stable. The molar ratios of leucyl and methionyl residues in individual proteins and in total protein were calculated by comparing their 3H/35S ratios to that of a protein with known amino acid composition. These values varied from 1.7 to 12. The half-lives of the unstable proteins did not exhibit a correlation with protein molecular weights, isoelectric points, or leucine/methionine ratios. It may be significant, however, that 6 of the 10 most unstable proteins had low leucine/methionine ratios whereas only 11% of all proteins tested had similarly low ratios.

The magnitude of differential reactivity effects that result from spectral differences in different portions of a pressurized water (PWR) core is studied, and it is shown that these effects can be correlated very well with the local moderator history. The impact of these differences on physics parameters such as axial offset, isothermal moderator temperature coefficient, and differential control rod worth is shown to be significant for two PWRs of considerably different design.

The stability of tryptophan biosynthetic enzyme activities was examined in cultures of repressor-negative (trpR) strains of Escherichia coli K-12 incubated under conditions of nutrient starvation of chloramphenicol inhibition. The results show that four of the five activities examined are stable under most nongrowing conditions, whereas one activity, indoleglycerol phosphate (InGP) synthetase, carried by the trpC protein, is unstable under most conditions tested. Phosphoribosylanthranilate (PRA) isomerase activity, which is also carried by the trpC protein, is unstable during starvation for ammonium, cysteine, or sulfate but is stable under other nongrowing conditions where InGP synthetase is not. InGP synthetase activity but not PRA isomerase activity is also diminished about twofold in cultures using glycerol as a carbon-energy source. These results indicate that one or both activities of the trpC protein is specifically inactivated under several culture conditions. Experiments with antibodies to the trpC protein show that sulfate-starved and ammonium-starved cultures contain 20 to 40% less immunologically reactive trpC protein than unstarved cultures. This indicates that the trpC protein is probably partially degraded under these conditions. During recovery from sulfate starvation or ammonium starvation, cultures slowly regain normal levels of InGP synthetase and PRA isomerase activities, suggesting that inactivation may be reversible.

Purified preparations of the bifunctional, tryptophan biosynthetic enzyme N-(5'-phosphoribosyl)anthranilate isomerase/indole-3-glycerol phosphate synthase of Escherichia coli K12 contain five forms of the protein (molecular mass = 49 kDa) which can be separated by isoelectric focusing or two-dimensional gel electrophoresis. The stability of the enzyme and its different forms was studied in exponentially growing, ammonium-starved and energy-depleted cultures using a dual-labeling, pulse-chase method. Labeled enzyme was isolated by standard purification techniques and by immunoprecipitation from crude extracts. The results demonstrated the following: 1) post-translational modification occurred in vivo, 2) modification occurred to the same extent in growing and ammonium-starved cells, 3) modification was dependent on a carbon-energy source for the cells, and 4) the enzyme was not degraded in growing or nongrowing cells. The pulse-chase data also indicated that one form of the protein (band 1) was the precursor of three other forms (bands 3-5) and that the relative amount of one form (band 2) remained constant.

The adequacy of the Advanced Recycle Methodology Program for predicting isotopic ratios as a function of exposure is determined for a light water reactor of modern design. For the first time, the capabilities of a two-dimensional multigroup transport theory code have been applied to the calculation of isotopic parameters for a commercial U.S. pressurized water reactor.The calculated results are found to be in satisfactory agreement with benchmark measurements. However, captures in seem to be underpredicted when ENDF/B-IV (or ENDF/B-III) resonance parameters for the 1.056-eV resonance are used in the calculations.

Received April 18, 1973 The utility of projection operator formalisms for describing the dynamics of many-body systems is studied, and the compatibility of these formalisms with certain approximation schemes is evaluated in the light of known behavior of such systems. For simplicity the investigation is limited to the study of Brownian motion. Specifically, a memory kernel formalism and a kinetic equation formalism are compared for the calculation of the time evolution of the momentum autocorrelation function. Both perturbation expansions and averaged propagator approximations are investigated. The results from these studies suggest that the long-time behavior of the momentum autocorrelation function is sensitive to the long-range nature of the interparticle potential.

A clinical observation that patients with aortoiliac occlusive disease have a high aortic bifurcation serves as the stimulus for this study. A review of 100 consecutive abdominal aortograms revealed that patients with occlusive disease had an average bifurcation angle of 38 degrees and those with normal studies or aneurysmal disease had an average angle of 52 degrees. A flow visualization system was constructed to study shear stress using eight Pyrex models with varying bifurcation angles (20 to 90 degrees). The perfusate consisted of a 35% sucrose solution with anion-exchange beads to serve as flow particles. The bifurcations were photographed at three different flow rates. The length of each tracer particle was measured to determine its velocity. Velocity profile curves were constructed and shear stress calculated by the formula shear stress = mu x dv/dr, where mu is viscosity, v is velocity, and r is radius. Results of the study suggest that more acute angles cause a greater shear stress on the inner wall of the bifurcation and decreased shear stress on the outer wall. Two theories of atheroma predilection, high shear stress and low shear stress, are applied to patients with high acute aortic bifurcations.

The metabolism of 184 individual proteins was examined in exponentially growing Escherichia coli. Cells were labeled with [3H]leucine and [35S]methionine using either a pulse-chase or a continuous labeling method. Proteins were fractionated by two-dimensional gel electrophoresis and their stabilities relative to total protein were determined from the 3H/35S ratios. Forty-seven proteins appeared to be unstable and were probably either degraded or modified to electrophoretically distinct forms. The apparent half-lives of these proteins calculated from the pulse-chase data varied from 2.0 to 23 h. Twenty-seven proteins appeared to be products of post-translational modifications. One hundred fourteen proteins appeared to be stable. The molar ratios of leucyl and methionyl residues in individual proteins and in total protein were calculated by comparing their 3H/35S ratios to that of a protein with known amino acid composition. These values varied from 1.7 to 12. The half-lives of the unstable proteins did not exhibit a correlation with protein molecular weights, isoelectric points, or leucine/methionine ratios. It may be significant, however, that 6 of the 10 most unstable proteins had low leucine/methionine ratios whereas only 11% of all proteins tested had similarly low ratios.

Purified preparations of the bifunctional, tryptophan biosynthetic enzyme N-(5'-phosphoribosyl)anthranilate isomerase/indole-3-glycerol phosphate synthase of Escherichia coli K12 contain five forms of the protein (molecular mass = 49 kDa) which can be separated by isoelectric focusing or two-dimensional gel electrophoresis. The stability of the enzyme and its different forms was studied in exponentially growing, ammonium-starved and energy-depleted cultures using a dual-labeling, pulse-chase method. Labeled enzyme was isolated by standard purification techniques and by immunoprecipitation from crude extracts. The results demonstrated the following: 1) post-translational modification occurred in vivo, 2) modification occurred to the same extent in growing and ammonium-starved cells, 3) modification was dependent on a carbon-energy source for the cells, and 4) the enzyme was not degraded in growing or nongrowing cells. The pulse-chase data also indicated that one form of the protein (band 1) was the precursor of three other forms (bands 3-5) and that the relative amount of one form (band 2) remained constant.

The bifunctional tryptophan biosynthetic enzyme N-(5'-phosphoribosyl)anthranilate (PRA) isomerase/indole-3-glycerol phosphate (InGP) synthetase was purified from Escherichia coli cultures incubated under several conditions, some of which result in inactivation of the enzyme (starvation for ammonium or sulfate, chloramphenicol inhibition). Recovery of enzyme activity during purification from cultures incubated under inactivating conditions suggested that activity was restored or that an inhibitor was removed. In these preparations, the enzymatic properties were altered slightly but not in a manner that could account for inactivation. Each preparation exhibited one major protein species (Mr approximately 50,000) when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis but revealed several species during isoelectric focusing in the range of pH 5.5 to 6.0. Comparison of the distribution of species indicated that modification to the more acidic forms had occurred in growing and nongrowing cells but not during stationary phase, glucose starvation, or energy depletion. The results suggested that inactivation and modification had occurred in vivo by an energy-dependent process. Several metabolites tested did not inhibit the purified enzyme. Attempts to detect an inhibitor in crude cell extracts were also not successful.

The elution profiles of Asp-tRNA from unstarved and starved cultures of a relaxed-control (Rel-) strain of Escherichia coli were compared by reversed-phase chromatography. Methionine starvation results in the appearance of several additional species of Asp-tRNA which are not observed with starvation for leucine or histidine. By the criterion of cyanogen bromide-effected shifts in chromatographic elution position, a large portion of the tRNAAsp synthesized in methionine-starved cells lacks the normal Q nucleoside. By the same criterion, virtually all of the tRNAAsp from unstarved, leucine-starved, and histidine-starved cells contain Q. We conclude that methionine starvation prevents the formation of the norma Q nucleoside in Rel- E. coli.