Dissociation Events Research Articles

The kinetic equation for the chloride transport cycle of band 3. A 35Cl and 37Cl NMR study.

The nature of a transmembrane transport process depends largely on the identity of the reaction that is rate-limiting in the transport cycle. The one-for-one exchange of two chloride ions across the red cell membrane by band 3 can be decomposed into two component reactions: 1) the binding and dissociation of chloride at the transport site, and 2) the translocation of bound chloride across the membrane. The present work utilizes 35 Cl NMR and 37 Cl NMR to set lower limits on the rates of chloride binding and dissociation at the saturated inward- and outward-facing band 3 transport sites (greater than or equal to 10(5) events site-1 s-1 in all cases). At both 0-3 and 37 degrees C, the NMR data specify that chloride binding and dissociation at the saturated transport sites are not rate-limiting, indicating that translocation of bound chloride across the membrane is the slowest step in the overall transport cycle. Using these results, it is now possible to describe many features of the kinetic equation for the ping-pong transport cycle of band 3. This transport cycle can be decomposed into two half-reactions associated with the transport of two chloride ions in opposite directions across the membrane, where each half-reaction is composed of sequential binding, translocation, and dissociation events. One half-reaction contains the rate-limiting translocation event that controls the turnover of the transport cycle; in this half-reaction, translocation must be slower than binding and dissociation. The other half-reaction contains the non-rate-limiting translocation event that in principle could be faster than binding or dissociation. However, when the following sufficient (but not necessary) condition is satisfied, both translocation events are slower than binding and dissociation: if the non-rate-limiting translocation rate is within a factor of 10(2) (0-3 degrees C) or 2 (37 degrees C) of the overall turnover rate, then translocation is rate-limiting in each saturated half-reaction. Thus, even though chloride appears to migrate through a channel that leads from the transport site to solution, the results support a picture in which the binding, dissociation, and channel migration events are rapid compared to the translocation of bound chloride across the membrane. In this case, chloride binding to the transport site can be described by a simple dissociation constant (KD = kappa OFF/kappa ON) rather than by a Michaelis-Menten constant (KM = (kappa OFF + kappa TRANSLOCATION)/KAPPA ON).

Isolation of unliganded steroid receptor proteins by high-performance size-exclusion chromatography : An investigation of steroid-dependent structural alterations

The physicochemical properties of unliganded steroid receptor proteins remain largely unknown primarily due to receptor lability in the absence of specific ligand, especially during prolonged biochemical analyses. We have utilized high-performance size-exclusion chromatography (HPSEC) as a rapid means of investigating the structural properties of cytosolic estrogen receptor proteins in both the presence and absence of [ 3 H]estradiol-17β. Cytosols prepared from immature calf uteri were analyzed by HPSEC on an Altex TSK-3000 SW column (600 mm × 7.5 mm I.D.) either before or after incubation with 10 n M [ 3 H]estradiol-17β. Postcolumn detection of previously unliganded receptor was accomplished by incubation of fractions (0.38 ml) with 10 n M [ 3H]estradiol-17β for 2–18 h at 0°C. Receptor-bound steroid was separated from free steroid by incubation with small pellets of hydroxylapatite. Nonspecific binding of [ 3H]estradiol-17β in parallel fractions was estimated using an unlabelled competitor (diethylstilbestrol) specific for the estrogen receptor. In low ionic strengths the receptor exists as a single, relatively stable, large form (retention time 34 min). The elution properties of this receptor configuration do not depend on bound steroid ligand. Analysis of receptor at elevated ionic strengths in the presence and absence of steroid ligand suggests that the salt-induced dissociation of receptor components to smaller forms (retention time 47 min) may be partially steroid-dependent. Characterization of receptor in 6 M urea demonstrates the presence of intermediate-sized receptor components (retention time 36–38 min). Analyses of receptor in 6 M urea—0.4 M potassium chloride suggests an inhibition of the more extensive salt-induced dissociation event seen in 0.4 M potassium chloride alone. Furthermore, the intermediate-sized receptor forms seen under these conditions (retention time 41–42 min) are generated in a steroid-dependent manner. The preparation of different molecular forms of biologically active, unliganded estrogen receptor by HPSEC should help further our investigations into the molecular mechanism(s) by which steroid hormones exert their receptor-mediated effects on target cells.

Photofragmentation dynamics of formaldehyde: CO(v,J) distributions as a function of initial rovibronic state and isotopic substitution

Complete rotational distributions have been obtained for the CO produced following excitation of H2CO, HDCO, and D2CO near the S1 origin. The CO was detected by vacuum ultraviolet laser-induced fluorescence. The distributions show a remarkable amount of rotational excitation, peaking at J=42, 49, and 53 for H2CO, HDCO, and D2CO, respectively, with widths of 20–25 J units (FWHM). CO(v=1) from H2CO photolysis has nearly the same rotational distribution as CO(v=0). The population of CO(v=1) is 14%±5% as large as the population of CO(v=0), in good agreement with earlier measurements. Increased angular momentum of H2CO is only partially transferred to CO, giving slightly wider rotational distributions without changing the peak value. The rotational distributions are highly nonthermal, showing that energy randomization does not occur during the dissociation event. An approximate range of product impact parameters has been determined. The impact parameters are too large to be accounted for by forces along the directions of the C–H bonds. The hydrogen appears to be most strongly repelled by the charge distribution a fraction of an Å outside the carbon atom of the CO. The distribution of impact parameters and the internal energy of the hydrogen fragment apparently do not change significantly upon isotopic substitution. The absence of population in CO(J<20) confirms the identity of CO(J≳25) as the long-lived intermediate in formaldehyde photodissociation.

Multiple states of U3 RNA in Novikoff hepatoma nucleoli.

U3 RNA, a capped small nuclear RNA found thus far only in the nucleolus, has been implicated in the processing and/or transport of preribosomal RNA [Busch, H., Reddy, R., Rothblum, L., & Choi, Y. C. (1982) Annu. Rev. Biochem. 51, 617-654]. Tris(hydroxymethyl)aminomethane (Tris) (10 mM, pH 7.0) extracts of Novikoff hepatoma nucleoli, which contained about 80% of total nucleolar U3 RNA, were analyzed by sucrose density gradient centrifugation. Approximately 65% of the U3 RNA was bound to greater than 60S preribosomal ribonucleoprotein (RNP) particles, and about 15% sedimented at less than 20 S. The association between the 65% of U3 RNA that was bound to the preribosomal RNP particles was stable up to 55 degrees C. About 10% of U3 RNA was base paired to preribosomal RNA after deproteinization at 22 degrees C. The base-paired fraction of U3 RNA was released from the preribosomal RNA by heating to 45 degrees C or treating with 4 M urea. These results show that of the total nucleolar U3 RNP, (a) about 55% is bound to preribosomal RNP particles primarily by protein interactions, (b) about 10% is base paired to preribosomal RNA, (c) approximately 15% sedimented slowly and consisted presumably of free U3 RNP particles, and (d) the remaining 20% of U3 RNP was not extractable using 10 mM Tris buffer. On the basis of the different association states of U3 RNP particles, a model is proposed for the binding and dissociation events which take place between U3 RNP and preribosomal RNP particles.

Weak beam observation of dissociated dislocations in an L12ordered alloy deformed at the temperature of strength anomaly

Abstract The dissociation properties of dislocations resulting from the compression of polycrystalline Ni3Al at 620°C and 800°C have been studied using the weak beam technique. The dissociation of screw dislocations on {001} is confirmed. However, it is shown that the non-screw segments are neither dissociated in the same plane as the screw parts, nor are they dissociated by a pure glide process. It is shown in several-examples that climb has played an extensive role during the dissociation event, a fact which should have an importance in the interpretation of the anomalous temperature dependence of the strength of L12 ordered alloys.

Apparatus for measurements on the fragmentation of MeV molecular-ion beams

An apparatus for the study of the fragmentation of MeV molecular ions is described. The system permits high-resolution measurement of the joint distribution in energy and angle for fragment ions of selected charge and mass. Two movable detectors allow the study of spatial and temporal correlations among up to three fragments resulting from a single dissociation event. All experimental parameters are monitored, controlled, displayed, and recorded by a computer system that uses a linked network of four processors. Real-time, computer-generated color graphics are employed to give a visual rendition of the relation between the detector positions and the trajectories of outgoing fragments.

Dynamics of ethyl radical decomposition. II. Applicability of classical mechanics to large‐molecule unimolecular reaction dynamics

AbstractThe classical trajectory method is used to investigate the unimolecular dynamics of ethyl radical dissociation. It is found that chaotic trajectories need not be backward integrable to yield accurate lifetime, and product energy and angular momenta distributions. This allows the use of large numerical integration step sizes in trajectory calculations. The product energy and angular momenta distributions are independent of the ethyl radical lifetime, and are obtained after only 50 dissociation events. Differences between classical and quantal unimolecular dynamics are discussed, and a prognosis for future trajectory studies of large‐molecule unimolecular decompositions is given.

In vitro and in vivo inhibition of primary transcription of vesicular stomatitis virus by a defective interfering particle.

A unique defective interfering (DI) particle, generated by a heat-resistant (HR) mutant of Indiana serotype vesicular stomatitis virus, was capable of inhibiting primary transcription by heterologous New Jersey serotype virions. The correlation between this phenomenon and the lowering of viral yields from doubly infected cells was investigated by the construction of chimeric DI particles containing the HR DI particle genome with a thermolabile polymerase. At the nonpermissive temperature, these DI particles were unable to self-transcribe, inhibit virion primary transcription, or reduce virion yield, but were able to be replicated. These results suggested that self-transcription of the HR DI particle genome was a prerequisite for heterotypic interference, but not for its own replication. Inhibition of virion primary transcription by HR DI ribonucleocapsids was also observed in vitro. At low HR DI to virion ribonucleocapsid ratios, the extent of inhibition was concentration dependent, whereas at high ratios, the amount of inhibition was concentration independent, approaching a limiting maximum value. A speculative mathematical model, which quantitatively accounts for these data, is presented. According to this model, the higher affinity for polymerase molecules by the HR DI ribonucleocapsids is explained in terms of dissociation events during transcription, which are more frequent in the longer virion ribonucleocapsids.

Head-to-tail polymerization of microtubules in vitro

The kinetics of microtubule polymerization to steady-state and the ability of tubulin subunits to exchange with polymer at steady-state were examined to determine the applicability of the head-to-tail polymerization mechanism (Wegner, 1976) to microtubule assembly in vitro. Under conditions where self-nucleation was a rare event, tubulin was induced to polymerize by the addition of short microtubule fragments, and the kinetics of elongation were analyzed as a pseudofirst-order reaction. At steady-state, a trace amount of [ 3H]tubulin, prepared by labeling in vivo of chick brain protein, was added to polymerized microtubules and the kinetics of label uptake into polymer were monitored by a rapid centrifugal assay. The isotope exchange kinetics were analyzed according to a theoretical model previously applied to actin polymerization (Wegner, 1976) and extended for the case of microtubule polymerization. The rate of head-to-tail polymerization, expressed as the steady-state subunit flux, was 27·6 ± 7·6 per second at 37 °C. The head-to-tail parameter s, a measure of the efficiency of subunit flux, was 0·26 ± 0·07, indicating that four association and four dissociation events resulted in the flux of one subunit through the polymer at steady-state. The role of GTP in this mechanism of microtubule polymerization was examined by replacement of the nucleotide occupying the exchangeable binding site of tubulin with the non-hydrolyzable GTP analog guanosine 5′-(β,γ-methylene)triphosphate. It was found that the rate of steady-state flux was reduced by two orders of magnitude compared to tubulin polymerized with GTP. The head-to-tail parameter approached its limiting value of zero, indicating greatly reduced efficiency of subunit flux through the polymer in the presence of this analog. In summary, this study demonstrates that microtubules exhibit significant headto-tail polymerization in the presence of GTP and, in keeping with theoretical considerations, provides evidence that nucleotide hydrolysis is required for subunit flux through the polymer.

Multiphoton decomposition of hexafluoroacetone: Effects of pressure, fluence, wavelength, and temperature on the decomposition yield and C-13 and O-18 isotopic selectivity

The yield and isotopic selectivity of the infrared multiphoton induced decomposition of hexafluoroacetone have been studied in both parallel and focused beam geometries as a function of temperature, pressure, fluence, and photolysing wavelength. At low conversions, the chemical stoichiometry of the decomposition process is simple, CF3COCF3 nhν CO + 2CF3. Several potential interferences are discussed but it is concluded that the yield of CO directly reflects the yield of the primary dissociation event. Pressure effects are complex and not amenable to quantitative description, however, varying parameters such as fluence and photolysing wavelength lead to predictions which give a clear qualitative picture. In particular, it is possible to distinguish parametric variations due to spontaneous dissociation and dissociation dependent upon energy pooling. Isotopic selectivity provides a powerful insight in this area. Preliminary data on the effect of temperature have been obtained at constant incident fluence and number density. Varying temperature probes the contribution of initially excited species (hot-bands). Reducing the temperature has no effect on the blue-edge of the absorption band but leads to a narrowing on the red-edge which is more marked for parallel beam geometries. The form of the wavelength dependence is in agreement with theoretical predictions. The observed carbon-13 isotopic selectivities of the dissociation are in basic agreement with those predicted from spectroscopy in that αCO is much higher than αC2F6. Very high values for this selectivity have been observed approaching αCO=1000 under extreme conditions. The equivalent selectivities for oxygen-18 are small, α?2. The hexafluoroacetone molecule is demonstrated to be a good model for study of multiphoton events and the data presented offer a powerful basis set for theoretical treatment.

The effect of strain-induced band-gap narrowing on high concentration phosphorus diffusion in silicon

A strain effect has been found to explain the anomalous reduction in P diffusivity at surface concentrations greater than ∼4×1020 cm−3. It is shown that at diffusion temperatures, misfit-induced strain causes a reduction in the effective Si band gap up to ∼−130 meV at the solubility limit of P. This band-gap narrowing results in reduced P diffusivity through a relative reduction of P+V= pairs in the surface region. This complex is the dominant species for P transport at high P concentrations. The diffusion of P in the tail region is dominated by V− vacancies liberated during P+V= pair dissociation events. Thus, the tail diffusivity will likewise be reduced by band-gap narrowing in the surface region. These reductions in diffusivity can be by as much as a factor of 6 depending upon temperature and P doping.

Infrared photodissociation of fluorinated ethanes and ethylenes: Collisional effects in the multiple photon absorption process

The ir photodissociation of fluorinated ethanes and ethylenes produces vibrationally excited HF via collisionless molecular elimination. Monitoring of the time resolved HF fluorescence is used as a probe to examine the effects that collisions with inert gas molecules have on the ir photodissociation process. The addition of inert gas is shown to enhance the dissociation yield. Mechanisms which may be responsible for this enhancement are discussed. An example of consecutive dissociation events within the same laser pulse is presented. The energy absorbed in excess of the minimum required for dissociation, the absorption cross section, and the energy absorption rate of highly vibrationally excited vinyl fluoride are estimated.

Rapidity dispersion analysis of individual events in π+p interactions at 8 to 23 GeV/c

An analysis has been performed of the dispersion of the rapidity spectrum of particles in individual events from well-identified exclusive channels of π + p interactions at 8, 16 and 23 GeV/ c . Emphasis is placed on testing the effectiveness of the method for separating different reaction mechanisms as a function of energy and multiplicity. By comparing data with simulated events generated according to transverse momentum limited phase space, we attempt to distinguish kinematical from dynamical features of the dispersion plots. The dispersion method isolates pion and baryon dissociation events fairly cleanly for sufficiently small masses of the excited systems.

S-and t-channel helicity conservation of pion induced nucleon diffraction dissociation at 3.9 GeV/c and the implications of a two-component model

A study of schannel and t-channel helicity conservation has been carried out on a prism plot separated sample of 3.9 GeV/ c π ±p → π (N π) dd diffraction dissociation events. s-channel helicity conservation is found to be strongly violated while t-channel helicity conservation is found to be mildly violated. These results are similar to that found at 11.7 GeV/ c. We explain these violations on the basis of a “two-component model” of which the first component is a single exchange amplitude. We postulate that this amplitude is the generalization of the pomeron exchange amplitude which may conserve s-channel helicity. The second component of the model is a double exchange mechanism which strongly violates s-channel helicity. The predictions of this model agree with the experimental data.

Collision Induced Dissociation of Molecular Ions

Velocity vector distributions of the fragment ion products of the dissociative collisions of O2+, N2+, NO+, and N2O+ with He have been determined, using projectile-target relative kinetic energies which are one to three times the bond energy of the molecular ion. The most probable dissociation event produces a fragment ion whose velocity is very nearly the same as that of the original projectile ion. Fragment ions also appear at smaller velocities and larger scattering angles, and the importance of these features increase with increasing initial relative energy. Three models for the dissociation are discussed, and it is concluded that a version of the stripping model is most nearly consistent with the data.

Vibrational Energy Exchange of Highly Excited Anharmonic Oscillators

The probabilities of dissociation Pdis, excitation Pex, de-excitation Pde, and the associated average energy exchanges were calculated theoretically for a highly energized, anharmonic oscillator (I2 or Br2) colliding classically with an inert gas atom C through a Morse-type potential. The effect of varying mass and well depth was investigated. It was found that the effect of anharmonicity is to favor excitation and dissociation. However, the average ΔEv for these processes is about the same for harmonic and anharmonic oscillators and not much bigger than 1 to 2 RT. Pdis increases markedly with increasing mass. The well depth (V0) does not seem to have much effect in the range 0.004<V0/D0<0.14, where D0 is the bond dissociation energy. Exchange reactions are observed, but for shallow well depths they are rare events. From the results, it is possible to calculate λ, the probability of stabilizing an atom pair by a third-body collision. This turns out to be independent of temperature but depends strongly on the mass of the third body C. It is always in the range 0.1<λ<1.0 for the systems considered, and increases with increasing mass of C. For the dissociation process, it is found that the average dissociation event gives rise to a pair of atoms with very little excess energy (∼0.10 RT) above the dissociation barrier. This implies an activation energy for dissociation of the order of D0°—2RT in the Benson—Fueno theory and also a T—2 dependence for the rate of recombination of atoms.