Protein denaturation: a small-angle X-ray scattering study of the ensemble of unfolded states of cytochrome c

Solution X-ray scattering was used to study the equilibrium unfolding of cytochrome c as a function of guanidine hydrochloride concentration at neutral pH. The radius of gyration (Rg) shows a cooperative transition with increasing denaturant with a similar Cm to that observed with circular dichroism. However, the lack of an isoscattering point in the X-ray scattering patterns suggests the equilibrium unfolding is not simply a two-state process. Singular value decomposition (SVD) analysis was applied to the scattering patterns to determine the number of distinct scattering species. SVD analysis reveals the existence of three components, suggesting that at least three equilibrium states of the protein exist. A model was employed to determine the thermodynamic parameters and the scattering profiles of the three equilibrium states. These scattering profiles show that one state is native (N). The other two states (U1, U2) are unfolded, with U2 being fully unfolded and U1 having some residual structure. Using the thermodynamic parameters to calculate fractional populations, U1 is maximally populated at intermediate denaturant concentrations while U2 is maximally populated at high denaturant concentrations. It is likely that there is a multiplicity of denatured states with U1 and U2 representing an average of the denatured states populated at intermediate and high denaturant concentrations, respectively.     

Modelling business processes in steel fabrication

This paper outlines the utilization of the Structured Analysis and Design Technique (SADT) in developing a business process model as part of an IMI-LINK research programme. The project investigates the integration of design and construction through a demonstrator project using steel roof trusses. The SADT modelling technique is a graphical language-based methodology that generates activity models depicting, in a successively detailed manner, process activities and their relationships to each other. The paper highlights the problems and limitations encountered during knowledge acquisition process for the model, the modelling process, and outlines the research programme for the re-engineering of the fabrication process.     

Effect of Pressure Treatment on Actomyosin ATPases from Flying Fish and Sardine Muscles

The effect of pressure treatment (1 atm ～ 5000 atm) on flying fish and sardine actomyosin ATPases was studied from the standpoint of interaction between myosin and actin. The activities of actomyosin Mg-ATPases markedly decreased and those of the EDTA-ATPases rapidly increased with prolonged pressure treatment at 3000 atm and 5000 atm. Changes in activities of F-actin plus pressure-treated myosin Ca-ATPases showed results similar to those of pressure-treated actomyosin Ca-ATPases, while myosin plus pressure-treated F-actin resulted in decreased Mg-ATPase activity and increased EDTA-ATPase activity.     

Two-level spinal stenosis in minipigs. Hemodynamic effects of exercise

STUDY DESIGN: Twenty-two G?ttingen minipigs were trained to run on a treadmill. Two-level lumbar spinal stenosis was created in 12 pigs, 10 were unoperated control subjects. Blood flow of the spinal cord and nerve roots was determined with microspheres at rest, during exercise, and after exercise. OBJECTIVES: To study the effect of lumbar spinal stenosis and exercise on blood flow of spinal neural tissue. SUMMARY OF BACKGROUND DATA: Neurogenic claudication, the key symptom of lumbar spinal stenosis, may be caused by vascular impairment or mechanical distress of neural tissue during exercise. Experimental compression of the cauda equina causes reversible nerve root edema, stasis, blood flow decrease, and compromised neural function. The vascular pathophysiology of spinal stenosis during exercise has not been studied previously. METHODS: Pigs were trained daily for 3 months. Two-level 25% lumbar spinal stenosis was introduced by placement of stenosing bands around the dural sac. Neurologic function was monitored before surgery by evoked potentials and after surgery by the Tarlov score. Regional blood flow in lumbosacral neural tissue was measured 3 days after chronic catheterization using microspheres at rest, during exercise at 3 km/h for 15 minutes, and at rest 30 minutes after exercise. RESULTS: Blood flow of grey and white matter increased during exercise in both groups, with no differences between groups. slight hyperemia prevailed after exercise in white matter of the stenotic area but not in grey matter. Nerve root blood flow was largely unchanged in control subjects during exercise but was reduced in spinal stenosis at rest, further depressed during exercise, and normalized after exercise. Dural blood flow was elevated throughout. CONCLUSION: The study suggests that exercise-induced impairment of spinal nerve root blood flow plays a role in the pathophysiology of neurogenic claudication.     

Echocardiographic evaluation of patients with a left ventricular device

Limited donor heart availability is primarily responsible for the renewal of interest in mechanical left ventricular assist devices (LVADs) as a bridge to transplantation. Donor availability is unlikely to increase significantly in the near future. Experience to date has shown that many patients can be maintained long enough to undergo transplantation, and LVADs may be acceptable as alternate therapy in some who may not be candidates for transplantation. However, criteria for noninvasive evaluation of patients on LVADs have not been developed. In a prospective study using serial echocardiography, we found that aortic valve opening, aortic forward flow, nonlaminar flow in the left ventricle, and mismatch of Doppler derived cardiac output at the pulmonic valve and device output are associated with device malfunction. Echocardiography was diagnostic in five of six patients with clinical instability unrelated to the device. These findings suggest that echocardiography is helpful in the routine evaluation of patients on LVADs.     

Selection of tumor antigens as targets for immune attack using immunohistochemistry: II. Blood group-related antigens

The Vietnam Era Twin (VET) Registry includes 14,800 male twins born 1939-55 and in military service in 1964-75. A mailed health survey including the Jenkins Sleep Questionnaire was sent to 11,959 members and 8,870 (74.2%) provided responses on the frequency of sleep problems in the previous month. Prevalence of those experiencing conditions at least 1 day per month was 67.2% for waking often, 61.5% for waking tired/worn out, 48.1% for trouble falling asleep and 48.6% for awakening early. Ordinal logistic regression analysis was used to estimate sleep problems from demographic, behaviour and lifestyle characteristics, and morbid conditions. Black (vs. white) race, older age, church/religious group participation, social support, employment, cigarette smoking, light physical activity, and strenuous physical activity were associated with lower risk of one or more sleep problems. Eleven morbid conditions with a prevalence of 1% or more, coffee consumption, heavy alcohol consumption, and Framingham Type A behaviour pattern were associated with a higher risk of sleep problems. These analyses suggest that sleep problems may be one of the mechanisms relating reduced quality of life to many physical and behavioural characteristics. Fortunately, a number of the risk factors associated with sleep problems are lifestyle characteristics which, if modified, may reduce sleep problems.     

Effects of in utero and lactational exposure of the laboratory rat to 2,4,2'',4''- and 3,4,3'',4''-tetrachlorobiphenyl on dopamine function

We have developed a biochemical approach for identifying the components of cortical actin assembly sites in polarized yeast cells, based on a permeabilized cell assay that we established for actin assembly in vitro. Previous analysis indicated that an activity associated with the cell cortex promotes actin polymerization in the bud. After inactivation by a chemical treatment, this activity can be reconstituted back to the permeabilized cells from a cytoplasmic extract. Fractionation of the extract revealed that the reconstitution depends on two sequentially acting protein factors. Bee1, a cortical actin cytoskeletal protein with sequence homology to Wiskott-Aldrich syndrome protein, is required for the first step of the reconstitution. This finding, together with the severe defects in actin organization associated with the bee1 null mutation, indicates that Bee1 protein plays a direct role in controlling actin polymerization at the cell cortex. The factor that acts in the second step of the reconstitution has been identified by conventional chromatography. It is composed of a novel protein, Pca1. Sequence analysis suggests that Pca1 has the potential to interact with SH3 domain-containing proteins and phospholipids.     

The complex characteristics of crystallization of the Fe75Si15B10 glassy ribbon

The Fe₇₅Si₁₅B₁₀ glassy ribbon was examined in the course of various isothermal and dynamic heat treatments. The number, type and mechanism of formation of the crystallization products formed during two crystallization stages in this alloy were determined by differential scanning calorimetry (DSC), thermomagnetometry, transmission electron microscopy and X-ray diffraction measurements. It is concluded that the first DSC peak is due to the crystallization of α-Fe(Si) or Fe₃Si and the composite microcrystals containing Fe₃B cores with α-Fe(Si) envelopes. The metastable Fe₃B subsequently transforms into the stable Fe₂B in the second DSC peak. The remaining amorphous matrix crystallizes by the eutectic reaction also in the second DSC peak forming the Fe₃Si and Fe₂B eutectic structures. The apparent activation energy E ₁ ^⊛ decreases during the first crystallization stage from 480 kJ (g atom^-1), characteristic of the Johnson–Mehl–Avrami (JMA) nucleation-and-growth kinetics of the as-quenched sample, to 350 kJ (g atom)^-1 implying a dominance of the diffusion-controlled growth of iron in the already pre-nucleated sample. Simultaneously, the JMA exponent n ₁ decreases from 2.5 to 1.5. The apparent activation energy E ₂ ^⊛ depends on the temperature of the first crystallization stage. It decreases with increasing T _a from 384 to 282 kJ (g atom)^-1. This phenomenon was attributed to modification of the chemical composition of the remaining eutectic after the temperature dependent primary crystallization of Fe₃B within the composites. The value of n ₂ is 3. This revised version was published online in November 2006 with corrections to the Cover Date.