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Biomedical subjects

F W Smith

Publications and source records attributed to F W Smith.

At least 127 records · Page 7Linked to original sources
PubMed1987

Purification of the yeast PHR1 photolyase from an Escherichia coli overproducing strain and characterization of the intrinsic chromophores of the enzyme.

We have placed the PHR1 gene of Saccharomyces cerevisiae under the transcriptional and translational control of the tac expression cartridge. Under inducing conditions Escherichia coli cells harboring plasmids carrying this construct accumulate approximately 8% of total cellular protein as the Phr1 photolyase. Using a strain devoid of E. coli photolyase activity, we have obtained milligram quantities of the yeast enzyme at greater than 95% purity and have characterized the enzyme. Phr1 photolyase is a monomer in solution with an Mr of 60,000, has a turnover number of 0.7 dimers min-1 molecule-1 in vitro, exhibits absorbance maxima at lambda = 277 and 377 nm, and has a fluorescence excitation maximum at 390 nm and an emission maximum at 475 nm. The near UV absorbance peak is shown to reflect the contributions of two intrinsic chromophores which are noncovalently bound to the enzyme. Spectroscopic, fluorescence, and thin layer chromatographic studies indicate that one of these chromophores is 1,5-reduced FAD rather than 4a,5-reduced FAD as previously proposed (Iwatsuki, N., Joe, C. O., and Werbin, H. (1980) Biochemistry 19, 1172-1176), while the other chromophore has properties similar to the second chromophore of E. coli photolyase. The fact that yeast and E. coli photolyases are similar both with respect to amino acid sequence and chromophore composition provides strong evidence that the enzymes share a common action mechanism which may also be utilized by photolyases from other organisms throughout the phylogenetic tree.

Amino Acids↗
PubMed1987

Action mechanism of Escherichia coli DNA photolyase. I. Formation of the enzyme-substrate complex.

Escherichia coli DNA photolyase (photoreactivating enzyme) is a flavoprotein. The enzyme binds to DNA containing pyrimidine dimers in a light-independent step and, upon illumination with 300-600 nm radiation, catalyzes the photosensitized cleavage of the cyclobutane ring thus restoring the integrity of the DNA. We have studied the binding reaction using the techniques of nitrocellulose filter binding and flash photolysis. The enzyme binds to dimer-containing DNA with an association rate constant k1 estimated by two different methods to be 1.4 X 10(6) to 4.2 X 10(6) M-1 S-1. The dissociation of the enzyme from dimer-containing DNA displays biphasic kinetics; for the rapidly dissociating class of complexes k2 = 2-3 X 10(-2) S-1, while for the more slowly dissociating class k2 = 1.3 X 10(-3) to 6 X 10(-4) S-1. The equilibrium association constant KA, as determined by the nitrocellulose filter binding assay and the flash photolysis assay, was 4.7 X 10(7) to 6 X 10(7) M-1, in reasonable agreement with the values predicted from k1 and k2. From the dependence of the association constant on ionic strength we conclude that the enzyme contacts no more than two phosphodiester bonds upon binding; this strongly suggests that the pyrimidine dimer is the main structural determinant of specific photolyase-DNA interaction and that nonspecific ionic interactions do not contribute significantly to substrate binding.

Collodion↗
PubMed1987

Magnetic resonance imaging of midline brain tumors using inversion recovery sequences at 0.08 T (3.4 MHz).

The use of inversion recovery sequences to highlight intracranial tumors in children is illustrated. The effect of changing the inversion time (TI) to produce the best spatial resolution and to highlight the contrast resolution between different tumors and normal brain is analyzed. The normal appearances and clinical examples in the central nervous system are used to illustrate the options that are available using IR sequences. Variation of TI for providing a means of differentiating short T1 lesions from long T1 lesions is discussed, short TI sequences being best for demonstrating gliomas and astrocytomas whilst medium/long TI sequences are best for demonstrating vascular abnormalities and short T1 tumors. Inversion recovery imaging is considered to be an accurate alternative to spin-echo imaging as currently applied.

Adolescent↗
PubMed1987

An NMR study of lithium responding manic depressive patients.

Manic depressive patients on long-term prophylactic therapy with lithium carbonate are examined by nuclear magnetic resonance imaging of the brain. Their ventricular brain ratios and cerebral grey and white matter spin lattice relaxation times (T1) are within normal limits. The implication of these findings in the light of previous studies using other techniques is discussed.

Adult↗
PubMed1987

Nitrogen Enhancement of Phosphate Transport in Roots of Zea mays L. : I. Effects of Ammonium and Nitrate Pretreatment.

The effect of nitrogen status on phosphorous uptake and translocation was examined in 6-day-old dark-grown decapitated maize seedlings exposed to 25 micromolar phosphorous. Transfer to complete solutions containing 1 millimolar ammonium resulted in an increase in phosphorous uptake rate after 6 to 8 hours. The stimulus remained effective for at least 5.5 hours upon subsequent transfer to nitrogen-free solutions. Pretreatments for 16 hours with either nitrate or ammonium resulted in enhanced rates of subsequent phosphorous uptake and in enhanced translocation to the xylem of the exogenously supplied phosphorous. Both processes reached a plateau following pretreatment with 0.1 to 1.0 millimolar concentrations of either nitrogen ion. Further enhancement occurred with 10 millimolar nitrate, but not with 10 millimolar ammonium pretreatment. Although nitrogen pretreatments slightly increased the quantity of exogenous phosphorous retained in the root tissue, most of the extra phosphorous taken up by the nitrogen-pretreated seedlings was translocated to the xylem. The enhanced translocation, however, did not totally account for the increase in uptake implying a specific stimulation of the uptake process.

Journal Article↗
PubMed1987

Nitrogen Enhancement of Phosphate Transport in Roots of Zea mays L: II. Kinetic and Inhibitor Studies.

Exposure to 1 millimolar ammonium pretreatments increased V(max) for phosphorous uptake in dark-grown decapitated maize seedlings without a statistically measurable change in K(m). Sulfate uptake also was stimulated. The stimulation in phosphorous uptake due to ammonium pretreatment was greater in seedlings grown without phosphorous than in those grown with 25 micromolar phosphorous. The stimulus was not expressed unless the entire root system was pretreated with ammonium, and pretreatment of a part of the root system inhibited phosphorous uptake by the remaining part unless it also had been pretreated. Pretreatment with the amino acid analogs p-fluoro-dl-phenylalanine and l-azetidine-2-carboxylic acid (AZ) restricted phosphorous uptake in seedlings that were pretreated with ammonium and in those that were not, but the effect of ammonium pretreatment was not completely eliminated by the analogs. In general, translocation of the entering phosphorous was affected similarly to uptake by experimental treatments. Enhanced translocation, however, was not sufficient to account quantitatively for the increase in uptake, and an increased uptake was still evident when translocation was completely prevented by 50 micromolar AZ pretreatment.

Journal Article↗
PubMed1987

Nuclear magnetic resonance imaging and single photon emission tomography with radio-iodine labelled compounds in the diagnosis of dementia.

White matter lesions and T1 changes were identified using NMR and then compared between groups of patients suffering from dementia of the Alzheimer type (DAT), Multiple infarct dementia (MID) and normal controls. All DAT and MID patients were also imaged with a gamma camera using 123Iodo-n-isopropyl-amphetamine, a radiopharmaceutical whose uptake in the brain follows the regional blood flow. While NMR was not able to differentiate between DAT and MID, 19 out of 21 DAT patients compared to four out of 18 MID patients showed bilateral parietal lesions on IMP scans.

Adult↗
PubMed1987

Diagnosis of orbital myositis by nuclear magnetic resonance imaging.

Two cases of orbital myositis are reported in which precise identification of the involved muscle was possible with the use of surface coil nuclear magnetic resonance (NMR) imaging. A clinical response to oral steroids in both cases coincided with a marked reduction in the abnormal thickening of the muscles as seen on the NMR images.

Administration, Oral↗
PubMed1987

Nuclear magnetic resonance brain imaging in chronic schizophrenia.

Patients with chronic schizophrenia were examined by nuclear magnetic resonance imaging of the brain. Subgroups of the syndrome with high positive or high negative symptom scores and ventricular dilatation were compared with each other and with normal controls in respect of regional spin lattice relaxation time (T1) changes. Significant differences were not observed between the schizophrenic subgroups and controls but there were significant differences between the subgroups themselves. The presence of tardive dyskinesia was associated with increased T1 of the basal ganglia. The significance of these results is discussed in relation to findings using other techniques.

Adult↗
PubMed1986

Magnetic resonance tomography of the pelvis.

Because magnetic resonance tomography (MRT) has the ability to make cross-sectional images in three planes that have superior contrast between organs, muscle, fat, and bone, its use for the demonstration of pelvic disease shows considerable promise. The normal pelvic anatomy is clearly displayed because of the high contrast between fat and other soft tissues, tumors of the bladder, prostate, ovary, and uterus and because of their different proton relaxation times show high contrast with the normal tissues. MRT has the ability to demonstrate benign and malignant tumors of pelvic organs and to display the extent of local and distal spread. Preliminary observations of its application to the diagnosis of bladder, prostatic, ovarian, and uterine malignancy are discussed. The absence of ionizing radiation for the production of the images allows for the safe imaging of the fetus in utero. Although it is not suggested that MRT should replace ultrasound for the examination of pregnancy, it appears that it will be a useful technique for studying fetal growth and placental function.

Female↗
PubMed1986

Measurement of total lung water from nuclear magnetic resonance images.

An accurate, non-invasive means of measuring pulmonary water is desirable, for both clinical and research use. Nuclear magnetic resonance imaging is theoretically very suitable for this purpose. Nuclear magnetic resonance (NMR) spectroscopy was used to show that the water content of lung tissue could be predicted accurately from the intensity of signal resulting from a 90 degree saturation recovery sequence. The water contained in an image section may, therefore, be calculated by reference to a 100% water standard. Lung water was obtained by imaging the lung in continuous sections and summing the water contents of the component sections. The method performed well when applied to a sponge phantom, but underestimated by up to 30% in excised sheep lung. The total (vascular and extravascular) pulmonary water measured by NMR in six healthy volunteers was 292 g (SD 58 g) or 4.6 g/kg body weight, less than predicted by some other indirect methods and post-mortem values. A briefer examination comprising two axial sections at standardised levels was also devised. In 15 healthy volunteers the mean water content of a 1.6 cm-thick axial section through the right lung was 17.8 g at the sternal angle, and 23.3 g 5 cm caudally. In the left lung, the values were 16.4 g and 16.3 g, respectively.

Adult↗
PubMed1986

Technetium-99m HM-PAO stereoisomers as potential agents for imaging regional cerebral blood flow: human volunteer studies.

A total of nine normal volunteer subjects were studied with three forms of [99mTc] hexamethylpropyleneamineoxime (HM-PAO), a potential cerebral blood flow imaging agent. One, the d,l isomer, showed 4.1% uptake in the brain which remained constant over 8 hr. There was good differentiation between uptake in gray and white matter on tomographic slices. We propose that this agent may allow regional cerebral blood flow imaging to be performed on a routine basis.

Adult↗
PubMed1985

Binding of Escherichia coli DNA photolyase to UV-irradiated DNA.

Escherichia coli DNA photolyase is a flavoprotein which catalyzes the photomonomerization of pyrimidine dimers produced in DNA by UV irradiation. In vivo, the enzyme acts by a two-step mechanism: it binds to dimer-containing DNA in a light-independent reaction and upon exposure to 300-500-nm light breaks the cyclobutane ring and dissociates from the substrate. Using photolyase purified to homogeneity, we have investigated in vitro the first step of the reaction, DNA binding; enzyme-DNA complex formation was quantitated by the nitrocellulose filter binding assay. We find that the enzyme binds specifically to UV-irradiated DNA regardless of whether the DNA is in the superhelical, open circular, or linear form or whether the DNA is single or double stranded. The binding reaction is optimum at a NaCl concentration of 125 mM and at pH 7.5. Although photolyase is retained by the nitrocellulose filters with near 100% efficiency, the binding efficiency of a single enzyme-substrate complex is about 0.34. The complexes can be dissociated by exposing them to photoreactivating light either in solution or on the filter.

DNA, Bacterial↗
PubMed1985

A comparison of T1 measurements at 1.7 and 3.4 MHz in the diagnosis of prostatic carcinoma.

To compare the specificity of T1 measurement for the diagnosis of prostatic carcinoma in vivo, two groups of patients with clinically enlarged prostate glands have been imaged. One group comprising 25 patients was imaged at 1.7 MHz the other comprising 51 patients at 3.4 MHz. T1 measurements together with the gross anatomical appearances of the prostate gland were observed and compared with the clinical and histologic diagnoses. At 1.7 MHz it was found that the T1 measurement of carcinoma of the prostate and benign hypertrophy of the prostate fell into two distinct ranges with no overlap between the two conditions. Three false-positive diagnoses of carcinoma were made in patients with prostatitis where the T1 values fell in the upper part of the carcinoma range. T1 measurement in the group studied at 3.4 MHz demonstrated no specific value for either condition and diagnosis of malignancy had to be made from the morphologic appearances of the gland. Morphologically benign hypertrophy appeared homogeneous while carcinoma appeared heterogeneous with small areas of increased T1 in the malignant part of the gland.

False Positive Reactions↗