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Biomedical subjects

J A Bell

Publications and source records attributed to J A Bell.

At least 19 recordsLinked to original sources
PubMed1992

Dissection of helix capping in T4 lysozyme by structural and thermodynamic analysis of six amino acid substitutions at Thr 59.

Threonine 59, a helix-capping residue at the amino terminus of the longest helix in T4 phage lysozyme, was substituted with valine, alanine, glycine, serine, asparagine, and aspartic acid. The valine, alanine, and glycine replacements were observed to be somewhat more destabilizing than serine, asparagine, and aspartic acid. The crystal structures of the different variants showed that changes in conformation occurred at the site of substitution, including Asp 61, which is nearby, as well as displacement of a solvent molecule that is hydrogen-bonded to the gamma-oxygen of Thr 59 in wild-type lysozyme. Neither the structures nor the stabilities of the mutant proteins support the hypothesis of Serrano and Fersht (1989) that glycine and alanine are better helix-capping residues than valine because a smaller-sized residue allows better hydration at the end of the helix. In the aspartic acid and asparagine replacements the substituted side chains form hydrogen bonds with the end of the helix, as does threonine and serine at this position. In contrast, however, the Asp and Asn side chains also make unusually close contacts with carbon atoms in Asp 61. This suggests a structural basis for the heretofore puzzling observations that asparagine is more frequently observed as a helix-capping residue than threonine [Richardson, J. S., & Richardson, D. C. (1988) Science 240, 1648-1652] yet Thr----Asn replacements at N-cap positions in barnase were found to be destabilizing [Serrano, L., & Fersht, A. R. (1989) Nature 342, 296-299].(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acids↗
PubMed1992

Theory of chaperonin action: inertial model for enhancement of prokaryotic Rubisco assembly.

We have performed a computational simulation of the aggregation and chaperonin-dependent reconstitution of dimeric prokaryotic ribulose bisphosphate carboxylase/oxygenase (Rubisco), based on the data of P. Goloubinoff et al. (1989, Nature 342, 884-889) and P. V. Viitanen et al. (1990, Biochemistry 29, 5665-5671). The aggregation is simulated by a set of 12 differential equations representing the aggregation of the Rubisco folding intermediate, Rubisco-I, with itself and with aggregates of Rubisco-I, leading up to dodecamers. Four rate constants, applying to forward or reverse steps in the aggregation process, were included. Optimal values for these constants were determined using the ellipsoid algorithm as implemented by one of us (Ecker, J.G. & Kupferschmid, M., 1988, Introduction to Operations Research, Wiley, New York, pp. 315-322). Intensive exploration of simpler aggregation models did not identify an alternative that could simulate the data as well as this one. The activity of the chaperonin in this system was simulated by using this aggregation model, combined with a model similar to that proposed by Goloubinoff et al. (1989). The model assumes that the chaperonin can bind the folding intermediate rapidly, and that the chaperonin complex releases the Rubisco molecule slowly, permitting time for its spontaneous folding while interacting with the chaperonin. This is followed by self-association of the folded Rubisco monomer to yield the active dimeric Rubisco. A modification of the model that simulates temperature effects was also constructed. The most important results we obtained indicate that the chaperonin-dependent reconstitution of Rubisco can be simulated adequately without invoking any catalysis of folding by the chaperonin. In addition, the simulations predict values for the association rate constant of Rubisco-I with the chaperonin, and other variables, that are subject to experimental verification.

Chaperonins↗
PubMed1992

p53 protein expression in human breast carcinoma: relationship to expression of epidermal growth factor receptor, c-erbB-2 protein overexpression, and oestrogen receptor.

The expression of p53 protein, oestrogen receptor protein, epidermal growth factor receptor (EGFR) and overexpression of the c-erbB-2 oncoprotein was examined in a series of 149 primary symptomatic breast carcinomas. Expression of p53 was present in 62 of 146 cases (42.5%) of the invasive carcinoma and one of three cases (33.3%) of ductal carcinoma in situ (DCIS) examined. Statistical associations of tumour oestrogen receptor positivity and lack of p53 protein expression, chi 2 = 19.78 (d.f. = 1), P less than 0.001, positive tumour p53 status and poor tumour grade; chi 2 = 14.1 (d.f. = 2), P less than 0.001, EGFR expression chi 2 = 7.07, (d.f. = 1), P less than 0.01 and tumour c-erbB-2 protein overexpression; chi 2 = 4.61 (d.f. = 1), P = 0.032 were identified. Expression of p53 is rare in invasive lobular carcinoma of classical type (8.3% of cases examined) in contrast to other common types of mammary carcinoma. Non-significant trends of p53 protein expression and increased regional tumour recurrence; chi 2 = 3.20 (d.f. = 1), P = 0.074 and also poorer patient survival; chi 2 = 3.76 (d.f. = 1), P = 0.053 were identified. p53 protein expression is a common event in human breast cancer and is present in both DCIS and invasive mammary carcinoma. Abnormal expression of p53 protein is a feature of both in situ and invasive breast carcinoma, implying that the abnormal p53 protein expression may be implicated in the early stages of mammary carcinoma progression.

Biomarkers, Tumor↗
PubMed1992

Ki67 immunoreactivity in breast carcinoma: relationships to prognostic variables and short term survival.

Immunoreactivity of the monoclonal antibody Ki67, which recognizes an antigen expressed in cells active in the cell cycle, has been investigated by immunocytochemistry in a series of 67 primary breast cancers. The percentage of tumour cell nuclei stained by Ki67 (labelling index) was related to tumour histological grade, mitotic frequency, oestrogen receptor status and tumour type. No correlation was found with patient age, tumour size or lymph node stage. A high Ki67 labelling index was significantly associated with diminished patient survival and disease-free interval, which demonstrates an important role for this monoclonal antibody as a prognostic marker in breast cancer.

Antibodies, Monoclonal↗
PubMed1991

Comparison of the crystal structure of bacteriophage T4 lysozyme at low, medium, and high ionic strengths.

Crystals of bacteriophage T4 lysozyme used for structural studies are routinely grown from concentrated phosphate solutions. It has been found that crystals in the same space group can also be grown from solutions containing 0.05 M imidazole chloride, 0.4 M sodium choride, and 30% polyethylene glycol 3500. These crystals, in addition, can also be equilibrated with a similar mother liquor in which the sodium chloride concentration is reduced to 0.025 M. The availability of these three crystal variants has permitted the structure of T4 lysozyme to be compared at low, medium, and high ionic strength. At the same time the X-ray structure of phage T4 lysozyme crystallized from phosphate solutions has been further refined against a new and improved X-ray diffraction data set. The structures of T4 lysozyme in the crystals grown with polyethylene glycol as a precipitant, regardless of the sodium chloride concentration, were very similar to the structure in crystals grown from concentrated phosphate solutions. The main differences are related to the formation of mixed disulfides between cysteine residues 54 and 97 and 2-mercaptoethanol, rather than to the differences in the salt concentration in the crystal mother liquor. Formation of the mixed disulfide at residue 54 resulted in the displacement of Arg-52 and the disruption of the salt bridge between this residue and Glu-62. Other than this change, no obvious alterations in existing salt bridges in T4 lysozyme were observed. Neither did the reduction in the ionic strength of the mother liquor result in the formation of new salt bridge interactions. These results are consistent with the ideas that a crystal structure determined at high salt concentrations is a good representation of the structure at lower ionic strengths, and that models of electrostatic interactions in proteins that are based on crystal structures determined at high salt concentrations are likely to be relevant at physiological ionic strengths.

Crystallization↗
PubMed1991

Low vision aids--is our service cost effective?

A questionnaire and telephone survey was carried out on a Scottish population of patients with impaired vision, in order to ascertain the proportion of patients who gain benefit from Low Vision Aids (LVA) and to determine the number of LVAs which are retained but unused. One third of the patients who answered the questionnaire never use their LVAs, and one half were not satisfied with the service provided. A cost analysis indicated that approximately (pounds) 8,000 worth of LVAs are neither used nor returned each year to a single LVA service. The patients' ages, diagnoses, and visual acuities were related to the compliance rate. It appears that increasing age and decreasing visual acuity may be factors which decrease compliance. However none of the factors analysed could be used as a reliable predictor of patient satisfaction or of eventual benefit. Other health services which provide intensive training in the use of LVAs reportedly achieve a higher level of compliance. We conclude that our present service could probably be improved by the employment of additional staff specifically trained to teach patients how to make best use of the LVAs provided.

Adolescent↗
PubMed1991

Plasma viscosity or erythrocyte sedimentation rate in the diagnosis of giant cell arteritis?

Plasma viscosity (PV) has replaced the erythrocyte sedimentation rate (ESR) as a routine laboratory test in many hospitals. The finding of a normal PV but raised ESR in a case of biopsy proved giant cell arteritis (GCA) cast doubt on this substitution in cases of suspected GCA. To assess the equivalence of PV and ESR in the diagnosis of this disease 40 suspected cases were prospectively investigated with both tests. The correlation between the two tests was good (r = 0.742, p less than 0.0001). The substitution of one test for the other would appear to be justified in most cases of suspected GCA. In the presence of biopsy proved disease, however, the PV and ESR each produced 13.3% false negatives. These occurred both in combination with and independently of the other test showing that, when in error, the two tests may not be equivalent. In cases of doubt the performing of both PV and ESR tests together improves but does not achieve complete diagnostic accuracy. Clinical judgment based on careful assessment of all available symptoms and signs must remain the foundation of diagnosis.

Aged↗
PubMed1991

Electrophysiological and binding studies on intact NCB-20 cells suggest presence of a low affinity sigma receptor.

Whole cell voltage clamp studies were performed on NCB-20 cells to examine physiological responses to drugs possessing affinities for sigma receptors. Those drugs [haloperidol, alpha-(4-fluoro-phenyl)-4-(5-fluoro-2-pyrimidinyl)-1-piperazinebutano l (BMY-14802), pentazocine, N-allylnormetazocine (SKF-10047), 3-(3-hydroxyphenyl)-N-(1-propyl)piperidine (3-PPP), phencyclidine, 1-[1-(2-thienyl)cyclohexyl]piperidine (TCP), (+)-5-methyl-10,11-dihydro-5H-dibenzo-[a,d]cyclohepten-5,10-imine maleate (MK-801)] caused an apparent inward current, which was due to blockade of a tonic, outward potassium current. The rank order of drug potencies in producing this effect generally resembled the rank orders of sigma-receptor affinities for the drugs, except that a reverse stereoselectivity was observed for several drugs. [3H](+)-SKF-10047 labeled two sites in intact NCB-20 cells (Kd = 49 nM, Bmax = 1.0 pmol/mg protein and Kd = 9.6 microM, Bmax = 69 pmol/mg protein). The high affinity site was similar pharmacologically to the sigma receptor assayed in membrane fragments from NCB-20 cells. However, the low affinity site showed a slightly different profile, highlighted by a reverse stereoselectivity. The rank order of drug potencies was as follows at the low affinity site: haloperidol greater than BMY-14802 greater than (-)-pentazocine greater than (+)-pentazocine greater than (-)-SKF-10047 greater than (-)-3-PPP greater than (+)-SKF-10047 greater than (+)-3-PPP greater than phencyclidine greater than TCP greater than MK-801.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

Evaluation of Campylobacter jejuni colonization of the domestic ferret intestine as a model of proliferative colitis.

Forty 3- to 17-week old domestic ferrets, including 2 gnotobiotes, were inoculated orally and/or rectally with 10(6) to 10(9) colony-forming units of 1 or more of 4 strains of Campylobacter jejuni, 3 of mink and 1 of human origin. Feeding or gavage of any of the 4 strains, in milk or broth, with or without preinoculation sodium bicarbonate treatment to neutralize stomach acid, induced colonization in 38/40 ferrets; diarrhea lasted 2 to 4 days in conventional kits, 6 days in gnotobiotes. Bacteremia was detected in 4 of 18 tested, 2 to 5 days after inoculation. Two strains caused no more severe disease or prolonged colonization after 3 serial IV passages in kits than they did before passage. Multiple inoculations with a given strain resulted in progressively briefer colonization and milder disease, but subsequent inoculation with a different strain induced colonization and gastrointestinal disease similar to a primary infection. Five kits inoculated rectally after 4 previous homologous inoculations were resistant to colonization as well as to disease. Agglutinin titers of ferrets inoculated orally or rectally once were low or undetectable, but increased in response to repeated inoculation. Pretreatment with a 1% formalin enema caused mild colon irritation without clinical or histologic evidence of proliferative colitis in ferrets concurrently inoculated orally and/or rectally, whether or not they had preexisting antibodies to any strain of C jejuni. Histologic examination of tissues revealed leukocytic infiltration of intestinal lamina propria in 29 of 35 infected kits and 5 of 8 noninfected controls, and cryptosporidiosis in 5 infected kits plus 1 control.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

Regulation of the NMDA receptor by redox phenomena: inhibitory role of ascorbate.

Redox phenomena seem to modulate activity of the N-methyl-D-aspartate receptor. Some reductants (ascorbate, hydroquinone) inhibit, while others (dithiothreitol, mercaptoethanol, penicillamine) potentiate NMDA receptor function. Ascorbate inhibits binding of [3H]glutamate and [3H]thienylcycohexylpiperidine to the NMDA receptor complex, and impedes NMDA-gated currents in isolated neurons; dithiothreitol-like reductants enhance NMDA-induced currents. The ability of reductants to alter function of the NMDA receptor is abolished by oxidation.

Animals↗
PubMed1990

Lack of detectable blood groups in domestic ferrets: implications for transfusion.

Evidence of blood groups in domestic ferrets was sought by testing serum samples for naturally acquired or experimentally induced erythrocyte antibodies. All sera were tested for ability to cause direct agglutination, antiglobulin-enhanced (Coombs test) agglutination, or lysis. Examination of 212 randomly paired combinations of ferret serum and erythrocytes produced no evidence of naturally acquired blood group antibodies. Six pairs of ferrets were reciprocally transfused twice, 34 days apart, with 6-ml quantities of anticoagulated blood. All were tested 21 days after the first transfusion, as well as 10 and 30 days after the second transfusion; erythrocyte antibodies were not detected. Four additional pairs of ferrets were reciprocally inoculated SC with a series of six 1.25-ml quantities of blood in Alsever solution, administered over a 3-week period, and tested 8 days after the last injection; again, erythrocyte antibodies could not be detected. These observations suggest that blood groups of the kind in human beings and other mammals either do not exist in domestic ferrets or represent antigen systems too weak to elicit measurable responses under the reported conditions. It appears, therefore, that transfusion in this species poses little clinical risk, even without crossmatching.

Agglutination Tests↗
PubMed1990

Clonidine attenuates increased brain glucose metabolism during naloxone-precipitated morphine withdrawal.

The effect of two doses of clonidine on regional cerebral metabolic rates for glucose were measured during morphine withdrawal in rats. In the first study, 0 or 200 micrograms/kg clonidine was administered to rats subjected to naloxone-precipitated morphine withdrawal (naloxone, 0.5 mg/kg, s.c.), and to non-dependent control rats. In a second study of similar design, 0 or 20 micrograms/kg clonidine were administered. Withdrawal signs in rats subjected to naloxone-precipitated morphine withdrawal and receiving 0, 20 or 200 micrograms/kg clonidine were also assessed. Naloxone-precipitated morphine withdrawal stimulated regional cerebral metabolic rates for glucose (59 of 83 regions in study no. 1; 73 of 83 regions in study no. 2). At 200 micrograms/kg, clonidine attenuated this effect (33 of 59 regions). Although 200 micrograms/kg clonidine directly suppressed regional cerebral metabolic rates for glucose in many regions (significant main effect of clonidine), it attenuated the naloxone-precipitated morphine withdrawal effect specifically in the lateral septal nucleus, medial habenula, subiculum and gracile nucleus (significant interactions between clonidine and morphine withdrawal). The 20 micrograms/kg dose of clonidine had no statistically significant effect. In behavioral experiments, both doses of clonidine diminished withdrawal in that there was no diarrhea, fewer wet-dog shakes and less abnormal posturing. However, locomotion, grooming and jumping were increased by clonidine. Most of these effects were statistically significant only with the 200 micrograms/kg dose. The results of these studies show that clonidine reduces morphine withdrawal-induced increases in regional cerebral metabolic rates for glucose in many brain regions, irrespective of the distribution of alpha 2-adrenoceptors. Although clonidine has been thought to ameliorate morphine withdrawal by actions primarily at the locus coeruleus and central amygdala, it may play a major role in other regions as well.

Animals↗