pH dependence of the stability of barstar to chemical and thermal denaturation.

Equilibrium unfolding of barstar with guanidine hydrochloride (GdnHCl) and urea as denaturants as well as thermal unfolding have been carried out as a function of pH using fluorescence, far-UV and near-UV CD, and absorbance as probes. Both GdnHCl-induced and urea-induced denaturation studies at pH 7 show that barstar unfolds through a two-state F<->U mechanism and yields identical values for delta GU, the free energy difference between the fully folded (F) and unfolded (U) forms, of 5.0 +/- 0.5 kcal.mol-1 at 25 degrees C. Thermal denaturation of barstar also follows a two-state F<->U unfolding transition at pH 7, and the value of delta GU at 25 degrees C is similar to that obtained from chemical denaturation. The pH dependence of denaturation by GdnHCl is complex. The Cm value (midpoint of the unfolding transition) has been used as an index for stability in the pH range 2-10, because barstar does not unfold through a two-state transition on denaturation by GdnHCl at all pH values studied. Stability is maximum at pH 2-3, where barstar exists in a molten globule-like form that forms a large soluble oligomer. The stability decreases with an increase in pH to 5, the isoelectric pH of the protein. Above pH 5, the stability increases as the pH is raised to 7. Above pH 8, it again decreases as the pH is raised to 10. The decrease in stability from pH 7 to 5 in wild-type (wt) barstar, which is shown to be characterized by an apparent pKa of 6.2 +/- 0.2, is not observed in H17Q, a His 17-->Gln 17 mutant form of barstar. This decrease in stability has therefore been correlated with the protonation of His 17 in barstar. The decrease in stability beyond pH 8 in wt barstar, which is characterized by an apparent pKa of 9.2 +/- 0.2, is not detected in BSCCAA, the Cys 40 Cys 82-->Ala 40 Ala 82 double mutant form of barstar. Thus, this decrease in stability has been correlated with the deprotonation of at least one of the two cysteines present in wt barstar. The increase in stability from pH 5 to 3 is characterized by an apparent pKa of 4.6 +/- 0.2 for wt barstar and BSCCAA, which is similar to the apparent pKa that characterizes the structural transition leading to the formation of the A form. The use of Cm as an index of stability has been supported by thermal denaturation studies.(ABSTRACT TRUNCATED AT 400 WORDS)     

Light-Induced Changes in Phosphorylation Status of Low Molecular Weight Wheat Nuclear Proteins

A large number of polypeptides were phosphorylated when in vitro protein phosphorylation was carried out in nuclei isolated from dark-grown seedlings. For studying the effect of light, dark-grown seedlings were exposed to light and the isolated nuclear proteins phosphorylated in vitro. Although 4 h of white light was sufficient to alter the phosphorylation status of at least two polypeptides of 19 and 17 kD but the effect of light was more pronounced after irradiation for 8 h or more, leading to virtual disappearance of a 19 kD and emergence of a 17 kD phosphopolypeptide. Studies using norflurazon, a bleaching herbicide, indicate that, in addition to 19 and 17 kD phosphopolypeptides, another 21 kD phosphopolypeptide may be involved in the de-etiolation process. However, the nature of the photoreceptor involved in these light-induced changes in nuclear protein phosphorylation remains to be established.     

Differentiation in explants from mature leguminous trees

Stem and petiole explants, obtained from mature trees, ofAlbizzia lebbeck,Cassia fistula andC.siamea callused and differentiated shoot-buds and later shoots on B5 medium supplemented with either 0.5 mg/l IAA + 1 mg/l BAP or BM + 2 mg/l NAA + 0.5 mg/l BAP. The stem explants were more responsive than the petiole explants. InA.lebbeck, the IAA substituted medium favoured differentiation from both types of explants. However, inC.fistula, the type of explants rather than the medium composition had an overriding influence on shoot differentiation since those from petiole hardly responded in either medium. It has been possible to obtain plantlets from bothA.lebbeck andC.fistula under conditions conducive to rooting. Plantlets ofA.lebbeck have also been successfully transferred to the field.     

Sensory innervation of the canine esophagus, stomach, and duodenum.

The sensory innervation of the postpharyngeal foregut was investigated by injecting the enzyme horseradish peroxidase (HRP) into the walls of the esophagus, stomach, or duodenum. The transported HRP was identified histochemically, labeled neurons in the spinal and vagal ganglia were counted, and the results were plotted using an SAS statistical program. The spinal sensory fields of each viscus were defined using three determinations: craniocaudal extent, principal innervation field, and peak innervation field. The data revealed that innervation fields are craniocaudally extensive, the sensory field of each viscus overlaps significantly with its neighbor, yet each viscus can be characterized by a field of peak innervation density. Craniocaudal innervation of the esophagus spans as many as 22-23 paired spinal ganglia (C1-L2). There are two peak innervation fields for the cervical (C2-C6 and T2-T4) and for the thoracic (T2-T4 and T8-T12) sectors of the esophagus. The sensory innervation of the stomach extends craniocaudally over as many as 25 paired spinal ganglia (C2-L5). The peak innervation field of the stomach spans a large area comprising the cranial, middle, and the immediately adjoining caudal thoracic ganglia (T2-T10). The duodenum is innervated craniocaudally by as many as 15 paired thoracolumbar ganglia (T2-L3). Peak innervation originates in the middle and caudal thoracic ganglia and cranial lumbar (T6-L1) ganglia. There is a recognizable viscerotopic organization in the sensory innervation of the postpharyngeal foregut; successively more caudal sectors of this region of the alimentary canal are supplied with sensory fibers from successively more caudal spinal dorsal root ganglia. Vagal afferent innervation of the esophagus, stomach, and duodenum is bilateral and originates predominantly, but not exclusively, from vast numbers of neurons in the nodose (distal) ganglia. The esophagus is innervated bilaterally and more abundantly by jugular (proximal) ganglia neurons than is either the stomach or duodenum. The physiological significance of the findings are discussed in relation to the phenomena of visceral pain and referred pain.     

THE EFFECT OF DIETARY HISTIDINE LEVEL ON THE CARNOSINE CONCENTRATION OF RAT OLFACTORY BULBS

Young adult male rats were fed purified diets supplying the maintenance level of the essential amino acids or the same diet devoid of histidine. Animals were sacrificed after 2, 4, 6 and 8 weeks on these diets and olfactory bulbs, whole brains and breast muscle removed for analysis of free histidine and histidinecontaining dipeptides. There was an immediate and sharp drop in the level of carnosine in the olfactory bulb of rats on the histidine-free diet. By 8 weeks only very small amounts of this dipeptide remained. The carnosine concentration in the olfactory bulbs of the rats receiving the maintenance level of histidine also decreased in comparison with the level maintained on the stock diet; this is believed to reflect the much reduced amount of histidine in the former as compared to the latter diet. Homocarnosine disappeared completely from whole brains of rats within 2 weeks on the histidine-free diet. Muscle carnosine decreased in both absolute terms and relative to the controls. Anserine was lower relative to the controls, but actually increased in absolute value. Histidine deficiency may be used to study the role of carnosine in olfactory function.     

Speckled SiO2@Au Core–Shell Particles as Surface Enhanced Raman Scattering Probes

Silica particles of ~800 nm size were functionalized using 3-amino propyl triethoxysilane molecules on which gold particles (~20 nm size) were deposited. The resulting particles appeared to form speckled SiO₂@Au core–shell particles. The surface roughness, along with hot spots, due to nanogaps between the gold nanoparticles was responsible for the enhancement of the Raman signal of crystal violet molecules by ~3.2?×?10⁷ and by ~1.42?×?10⁸ of single-wall carbon nanotubes. It has also been observed that the electromagnetic excitation near surface plasmon resonance (SPR) of core–shell particles is more effective than off resonance SPR excitation.     

Professional uncertainty and disempowerment responding to ethnic diversity in health care: a qualitative study

Background

While ethnic disparities in health and health care are increasing, evidence on how to enhance quality of care and reduce inequalities remains limited. Despite growth in the scope and application of guidelines on “cultural competence,” remarkably little is known about how practising health professionals experience and perceive their work with patients from diverse ethnic communities. Using cancer care as a clinical context, we aimed to explore this with a range of health professionals to inform interventions to enhance quality of care.

Methods and Findings

We conducted a qualitative study involving 18 focus groups with a purposeful sample of 106 health professionals of differing disciplines, in primary and secondary care settings, working with patient populations of varying ethnic diversity in the Midlands of the UK. Data were analysed by constant comparison and we undertook processes for validation of analysis. We found that, as they sought to offer appropriate care, health professionals wrestled with considerable uncertainty and apprehension in responding to the needs of patients of ethnicities different from their own. They emphasised their perceived ignorance about cultural difference and were anxious about being culturally inappropriate, causing affront, or appearing discriminatory or racist. Professionals'' ability to think and act flexibly or creatively faltered. Although trying to do their best, professionals'' uncertainty was disempowering, creating a disabling hesitancy and inertia in their practice. Most professionals sought and applied a knowledge-based cultural expertise approach to patients, though some identified the risk of engendering stereotypical expectations of patients. Professionals'' uncertainty and disempowerment had the potential to perpetuate each other, to the detriment of patient care.

Conclusions

This study suggests potential mechanisms by which health professionals may inadvertently contribute to ethnic disparities in health care. It identifies critical opportunities to empower health professionals to respond more effectively. Interventions should help professionals acknowledge their uncertainty and its potential to create inertia in their practice. A shift away from a cultural expertise model toward a greater focus on each patient as an individual may help.