Biomedical subjects
Publications and source records attributed to M Johnston.
Doctors, nurses and relatives involved with 30 recently aphasic stroke patients were asked to predict how the patient would perform on a comprehension test. Results show that not only do doctors, nurses and relatives underestimate the receptive disability of these patients, but they also illustrate a lack of agreement between health professionals. Implications for management are considered.
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The promoters of the highly expressed and stringently regulated GAL genes of Saccharomyces cerevisiae, are useful for expressing proteins in this organism. However, two problems complicate their use. First, because growth on glucose causes prolonged repression of GAL expression, cells are most rapidly induced after growth on nonfermentable carbon sources, conditions which usually support poor growth. Second, because the inducer of the GAL genes (galactose) also serves as a carbon source, the level of inducer is continually diminishing during growth of a Gal+ strain, which may lead to reduced GAL expression. To solve the first problem, we have employed strains that carry the reg1-501 mutation, which eliminates glucose repression of GAL expression. This gene has been shown to be located on the right arm of chromosome IV, distal but tightly linked to the TRP1 gene. We demonstrate that expression from GAL promoters is efficiently and rapidly induced in these reg1 strains by the addition of galactose to a culture growing in glucose medium. Levels of galactose as low as 0.02% can be used to obtain a 1500-fold induction of gene expression from GAL promoters in this strain. To surmount the second problem, we have used a gal1 mutant, deficient in the enzyme that catalyzes the first step of galactose utilization. We show that high levels of expression from GAL promoters are achieved rapidly in these mutants, for which galactose is a gratuitous inducer.(ABSTRACT TRUNCATED AT 250 WORDS)
The steady-state kinetic mechanism of highly purified bovine liver mitochondrial dihydroorotate dehydrogenase has been investigated. Initial velocity analysis using S-dihydroorotate and coenzyme Q6 revealed parallel-line, double-reciprocal plots, indicative of a ping-pong mechanism. The dead-end inhibition pattern with barbituric acid and the reactions with alternate cosubstrates methyl-S-dihydroorotate and menadione also point to a ping-pong mechanism. However, product orotate was found to be competitive with dihydroorotate and uncompetitive with Q6. These findings suggest that dihydroorotate dehydrogenase may follow a nonclassical, two-site ping-pong mechanism, typical of an enzyme that contains two non-overlapping and kinetically isolated substrate binding sites. That these two sites communicate by an intramolecular electron-transfer system involving FMN and perhaps an iron-sulfur center is also suggested by the kinetic behavior of the enzyme.
Dihydroorotates deuteriated at both C5 and C6 have been prepared and used to probe the mechanism of the bovine liver mitochondrial dihydroorotate dehydrogenase. Primary deuterium isotope effects on kcat are observed with both (6RS)-[5(S)-2H]- and (6RS)-[6-2H] dihydroorotates (3 and 6, respectively); these effects are maximal at low pH. At pH 6.6, DV = 3.4 for the C5-deuteriated dihydroorotate (3), and DV = 2.3 for the C6-deuteriated compound (6). The isotope effects approach unity at pH 8.8. Analysis of the pH dependence of the isotope effects on kcat reveals a shift in the rate-determining step of the enzyme mechanism as a function of pH. Dihydroorotate oxidation appears to require general base catalysis (pKB = 8.26); this step is completely rate-determining at low pH and isotopically sensitive. Reduction of the cosubstrate, coenzyme Q6, is rate-limiting at high pH and is isotopically insensitive; this step appears to require general acid catalysis (pKA = 8.42). The results of double isotope substitution studies and analysis for substrate isotope exchange with solvent point toward a concerted mechanism for oxidation of dihydroorotate. This finding serves to distinguish further the mammalian dehydrogenase from its parasitic cognate, which catalyzes a stepwise oxidation reaction [Pascal, R., & Walsh, C.T. (1984) Biochemistry 23, 2745].
The purpose of most screening and diagnostic tests is to initiate treatment when a problem is found and, otherwise, to allay anxiety. The extent to which the second of these objectives is met was studied in a prospective study of 179 pregnant women. Women who had amniocentesis to detect Down's Syndrome or were screened for a fetal neural tube defect had significantly lower anxiety levels in the third trimester of pregnancy than women who did not undergo these tests. Undergoing screening for a neural tube defect was also associated with a more positive attitude towards the pregnancy in the second trimester. These results suggest that the previously reported raised levels of anxiety in women undergoing such tests do not necessarily remain high for the duration of the pregnancy and, indeed, undergoing testing may serve to protect women against high levels of anxiety in the third trimester of pregnancy.
Mesophyll chloroplasts from sodium-deficient compared to normal plants of the C(4) species Kochia childsii and Amaranthus tricolor were found to have significantly less stacking in their grana. On the other hand, no marked difference of thylakoid arrangement between bundle sheath chloroplasts from sodium-deficient and normal plants of A. tricolor were observed.
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A two-year prospective study compared individual and group models of support for foster parents. Results showed greater foster parent satisfaction with the group model. An independent critique identified methodological flaws that would preclude further differentiation of the models and suggested that clinical impressions and trends in favor of the group model be given credibility. Operation of the group model is described, and reasons for the differences between clinical and experimental findings are discussed.
Perceived personal control has been found to be predictive of desirable outcomes in a variety of health care settings. It was hypothesized that it would also explain some of the variance in adaptation to and coping with physical disability and would therefore be predictive of recovery. A Recovery Locus of Control scale (RLOC) was developed, independent judges agreeing on five internal and four external items. The scale was shown to have good internal consistency when used with adults recently disabled by a stroke or wrist fracture. Patients' RLOC was assessed at the start of rehabilitation and recovery from physical disability was monitored using measures appropriate for each clinical group. In both groups, greater internality was associated with faster recovery. Results are discussed with respect to the likely mediating factors and the role of health professionals in facilitating internality.
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Records were reviewed of all patients with premature rupture of the membranes (PROM) at or less than 34 weeks estimated gestational age (EGA) who delivered at University Hospital of Jacksonville, Florida, during 1987. That year 132 patients were identified, 3.1% of all deliveries. The mean time from membrane rupture to delivery was three days, and the duration of PROM seven or more days in 9% of cases. Chorioamnionitis was diagnosed in 20% of the mothers. Delivery was by cesarean section in 30% of cases, twice the primary rate at University Hospital for 1987. Sixty-five percent of infants were male, and 13 males and four females of the 132 infants died before or after birth, the majority due to prematurity. Respiratory distress syndrome (RDS) was found in 35 infants and in 80% of these cases the membrane ruptured at 30 weeks or less. Intraventricular hemorrhage (IVH) was diagnosed in 8% of cases. Neonatal sepsis was a common diagnosis (41% of deliveries) with incidence being similar at all gestational ages. The length of membrane rupture was not statistically significant when compared with neonatal sepsis (P = 0.39).
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We constructed plasmids carrying the Escherichia coli proB gene that encodes gamma-glutamyl kinase, under the control of the yeast GAL1 promoter. This construction was carried out with both the wild-type proB+ gene and a mutant allele, proB74, that specifies an enzyme resistant to feedback inhibition by proline. Yeast pro1 mutants harboring these plasmids are proline prototrophs. We conclude that the pro1 mutation results in a deficiency in the gamma-glutamyl kinase activity in Saccharomyces cerevisiae. Expression of the proB74 allele in yeast resulted in enhanced resistance to the proline analogue L-azetidine-2-carboxylate and in a 2.4-fold elevation of the intracellular free proline levels. This result suggests that gamma-glutamyl kinase is the rate limiting step in proline biosynthesis in yeast.
A prerequisite to an informed decision to undergo any screening or diagnostic test is knowledge about such a test. This study describes the development of a self-administered questionnaire to measure knowledge of prenatal tests, for use in studies concerning the uptake of these tests. Both the reliability (internal and test-retest) and predictive validity are evaluated. Validity is assessed in two studies: first, comparison is made between three criterion groups (two with experience of pregnancy, one without); and second, scores on the questionnaire are compared before and after women have been provided with information about possible tests. The results show this to be both a reliable and valid measure of knowledge about prenatal tests. The results also highlight aspects of knowledge that are lacking. For example, a sizeable minority of women were uncertain or incorrect in identifying which tests they had undergone in a recent pregnancy.
A two-year (1984-1985) review of 3,764 questions appearing on Part I and Part II of the National Board of Medical Examiners (NBME) indicated 140 (4%) were transfusion medicine (TM)-related. Fewer questions on TM appeared on Part II than Part I (45 vs. 95, P less than 0.001). Unexpectedly, the lowest proportion of questions (excluding psychiatry) in the Part II examination was in the surgery section. Part I NBME appears to adequately reflect and assess the TM content of the medical school curricula, but the number and distribution of questions in Part II of the NBME does not reflect recent changes in the clinical transfusion medicine curricula of many United States medical schools.
The GAL4 protein of Saccharomyces cerevisiae binds to DNA upstream of each of six genes and stimulates their transcription. To locate regions of the protein responsible for these processes, we identified and characterized 88 gal4 mutations selected in vivo to reduce the ability to GAL4 protein to activate transcription. These mutations alter two regions of GAL4 protein: the DNA binding domain, and the transcription activation domain. Some mutations in the DNA binding domain that abolish the ability of GAL4 protein to bind to DNA in vitro change amino acid residues proposed to form a zinc finger, confirming that this structure is indeed involved in DNA binding. Four different amino acid changes in the zinc finger appear to reduce (but not abolish) the affinity of GAL4 protein for zinc ions, thereby identifying some of the amino acids involved in forming the zinc-binding structure. Several other mutations that abolish the DNA binding activity of the protein alter the 20 amino acids adjacent to the zinc finger, suggesting that these residues are part of the DNA binding domain. Two amino acid changes in the region adjacent to the zinc finger also appear to affect the ability of GAL4 protein to bind zinc ions, suggesting that this region of the protein can influence the structure of the zinc binding domain. The transcription activation domain of GAL4 protein is remarkably resistant to single amino acid changes: only 4 of the 42 mutations that alter this region of the protein are of the missense type. This observation is consistent with other lines of evidence that GAL4 protein possesses multiple transcription activation domains with unusual sequence flexibility.