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Biomedical subjects

J Weaver

Publications and source records attributed to J Weaver.

At least 19 recordsLinked to original sources
PubMed1992

Genetic analysis of a locus on the Bacteroides ovatus chromosome which contains xylan utilization genes.

Bacteroides ovatus, a gram-negative obligate anaerobe found in the human colon, can utilize xylan as a sole source of carbohydrate. Previously, a 3.8-kbp segment of B. ovatus chromosomal DNA, which contained genes encoding a xylanase (xylI) and a bifunctional xylosidase-arabinosidase (xsa), was cloned, and expression of the two genes was studied in Escherichia coli (T. Whitehead and R. Hespell, J. Bacteriol. 172:2408-2412, 1990). In the present study, we have used segments of the cloned region to construct insertional disruptions in the B. ovatus chromosomal locus containing these two genes. Analysis of these insertional mutants demonstrated that (i) xylI and xsa are probably part of the same operon, with xylI upstream of xsa, (ii) the true B. ovatus promoter was not cloned on the 3.5-kbp DNA fragment which expressed xylanase and xylosidase in E. coli, (iii) there is at least one gene upstream of xylI which could encode an arabinosidase, and (iv) xylosidase rather than xylanase may be a rate-limiting step in xylan utilization. Insertional mutations in the xylI-xsa locus reduced the rate of growth on xylan, but the concentration of residual sugars at the end of growth was the same as that with the wild type. Thus, a slower rate of growth on xylan was not accompanied by less extensive digestion of xylan. Mutants in which xylI had been disrupted still expressed some xylanase activity. This second activity was associated with membranes and produced xylose from xylan, whereas the xylI gene product partitioned primarily with the soluble fraction and produced xylobiose from xylan.

Bacteroides↗
PubMed1990

Two types of receptors for iron on mitochondria.

ATP, the major ligand for Fe in the reticulocyte's low-Mr chelatable pool, transfers Fe to a mitochondrial receptor from which the Fe is incorporated into haem. However, the utilization of Fe bound to this receptor for haem synthesis is slow in comparison with that bound to a second receptor that does not accept Fe from ATP, but does accept it from AMP. The major pathway by which Fe is delivered for haem synthesis may be through hydrolysis of cytosolic ATP-Fe to AMP-Fe.

Adenosine Monophosphate↗
PubMed1990

Mitochondria have Fe(III) receptors.

Recent work has provided new evidence that ATP is the major constituent of the low-Mr iron pool in the reticulocyte. The interaction of the iron complex of ATP with mitochondria was investigated in the present experiments. When ATP-Fe3+ was incubated with mitochondria, Fe3+, free of ATP, bound with high affinity to Fe3+ receptors on the mitochondria. The binding was saturable and reversible. Iron which was complexed to PPi, nitrilotriacetate, citrate, ADP and GTP also showed saturable binding to mitochondria; Fe3+ complexed to AMP bound non-specifically, as did Fe2+/ascorbate complexed to AMP bound non-specifically, as did Fe2+/ascorbate and Fe2+/dithionite.

Adenosine Triphosphate↗
PubMed1990

Iron bound to low MW ligands: interactions with mitochondria and cytosolic proteins.

The iron in the low MW pool of the cell is the precursor of iron in haem and is bound primarily to ATP. This precursor-product relationship suggested that reticulocytes might accumulate ATP-iron if their haem synthesis were blocked. However reticulocytes, treated with succinylacetone or rotenone and taking up iron from transferrin, accumulated iron in nonhaem cytosolic proteins and in mitochondria and not in the low MW pool. This was demonstrated by NMR and also by disrupting the cell with shear stress, separating the cytosol and pellet and fractionating the cytosol with ammonium sulfate. This constancy of the low MW iron pool in the face of blocked haem synthesis could not be explained by saturation of cytosolic ATP or by sluggish exchange of the low MW pool with other compartments. Rather, nonhaem cytosolic proteins and mitochondria appeared to have a higher affinity for iron and to exchange it rapidly with that in the low MW pool.

Animals↗
PubMed1990

Two pathways for iron uptake by guinea pig reticulocytes.

We have demonstrated that the intracellular processing of transferrin to effect iron removal involves two pathways, one sensitive to rotenone and the other not. We have also found that the effect of the rotenone is dependent on the transferrin concentration: iron uptake was suppressed with concentrations of transferrin in the micromolar range, and was not suppressed at physiologic concentrations of transferrin. Rotenone does not disturb transferrin's interaction with its extracellular receptor, indicating that its action must be intracellular. The following model is suggested: that separate pathways are entered by transferrin in the cell. The first pathway is preferentially utilized when transferrin is in short supply. It begins with an intracellular site which has a high affinity (and low capacity) for either iron or transferrin. The second pathway begins with an intracellular site which has a high capacity (but low affinity) for either iron or transferrin and is utilized when transferrin is in physiologic concentration (and the low-capacity, high-affinity site is saturated); the pathway it initiates is dominant when transferrin is abundant. We speculate that the high-affinity low-capacity pathway may serve to direct intracellular iron to sites which would be critically injured by iron excess.

Animals↗
PubMed1990

Iron binding to apotransferrin.

The classic analysis of metal transfer between ligands suggests that the metal-binding ligand (M-L1) and the uncomplexed ligand (L2) form a mixed complex (L2-M-L1), and that transfer is effected with the dissociation of this complex to L2-M and L1. Spectroscopic data suggested that such mixed complexes formed when pyrophosphate-Fe and acetohydroxamate-Fe were added to apotransferrin: the initial species had a different absorbance maximum than the final transferrin-iron complex. We now show that similar spectroscopic changes are seen when free ferrous iron or iron liganded to ATP, citrate or nitrilotriacetate are added to apotransferrin. The evolving spectrum on the addition of iron to apotransferrin may thus reflect iron binding per se rather than the formation of a mixed ligand complex.

Adenosine Triphosphate↗
PubMed1989

NMR studies of intracellular free calcium, free magnesium and sodium in the guinea pig reticulocyte and mature red cell.

During the maturation process reticulocytes lose their intracellular organelles and undergo changes in membrane lipid composition and ion transport properties. While several reports indicate differences in the levels of magnesium, sodium and calcium in reticulocytes and erythrocytes, controversy remains concerning the actual magnitude and direction of ionic alterations during reticulocyte maturation. One problem with all of these studies is that the techniques used are invasive and are limited to measuring only the total cell ion content. We have used 31P, 23Na and 19F nuclear magnetic resonance (NMR) spectroscopy to compare the intracellular free ion and phosphometabolite levels in guinea pig reticulocytes and mature red blood cells. In contrast to a sharply decreased concentration of ATP in erythrocytes in comparison to reticulocytes, the intracellular free magnesium, measured using 31P-NMR, was increased by about 65% upon maturation (150 mumol/l cell water in reticulocytes in comparison to 250 mumol/l cell water in erythrocytes). Sizeable but opposite changes in intracellular sodium (5.5 mumol/ml cells in reticulocytes vs. 8.5 mumol/ml cells in erythrocytes) and intracellular free calcium (99 nM vs. 31 nM in reticulocytes and mature red cells, respectively) were also observed, suggesting that alterations in the kinetics of membrane ion transport systems, accompanying changes in phospholipid and cholesterol content, occur during the process of red cell maturation. However, in contrast to dog red blood cells, there was no evidence for the presence of a Na+/Ca2+ exchanger in guinea pig reticulocytes or erythrocytes.

Animals↗
PubMed1989

Making choices.

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Arthritis↗
PubMed1989

Partially compensated hypoadrenalism presenting with persistent skin pigmentation.

A 33-year-old female presented in 1966 with striking pigmentation, typical of Addison's disease, and amenorrhea. Endocrine assessment then showed normal basal serum cortisol and urinary hydroxysteroid levels, but serum cortisol did not respond to stimulation with either exogenous ACTH or lysine vasopressin. Steroid replacement treatment was started. Treatment was discontinued by the patient on her own initiative and after some yr she was lost to follow-up. Reassessment in 1986 showed a pigmented patient who had continued in good health. She had a normal basal serum cortisol level with circadian variation. Plasma ACTH levels were high but showed diurnal rhythmicity and suppressed incompletely with 2 mg or 8 mg of dexamethasone/24 h. Plasma aldosterone levels were normal and showed appropriate postural changes, but plasma renin levels were high. This patient has an immunological profile of autoimmune disease with positive adrenal, thyroid microsomal and gastric parietal cell antibodies with a history of a premature menopause which may also be of autoimmune origin. She has been seen over a 20-yr period and despite her appearance still has no biochemical evidence of glucocorticoid or mineralocorticoid deficiency. It is suggested that the patient had compensated hypoadrenalism, with serum cortisol levels maintained in the normal range by high plasma ACTH levels and serum aldosterone levels maintained by high renin levels. The long term result of the high ACTH levels was increased skin pigmentation.

Adrenal Insufficiency↗
PubMed1989

An evaluation of volumes and concentrations of lidocaine in human inferior alveolar nerve block.

The purpose of this study was to evaluate, with the electric pulp tester, the anesthetic efficacy of 1.8 ml of 2% lidocaine with 1:100,000 epinephrine, 3.6 ml of 2% lidocaine with 1:200,000 epinephrine, and 1.8 ml of 4% lidocaine with 1:100,000 epinephrine in human inferior alveolar nerve block. Thirty subjects randomly received each of the solutions at three successive appointments. The first molar, canine, lateral incisor, and contralateral canine were tested with the pulp tester at various time intervals up to 55 min. Complete anesthesia was defined as an 80/80 reading with the pulp tester. No significant differences in anesthetic success or failure were found among the three solutions. Potential anesthetic problems (failure, noncontinuous anesthesia, slow onset, and short duration) occurred in 43 to 57% of the molars, in 43 to 60% of the canines, and in 57 to 80% of the lateral incisors. Complete anesthesia in the mandible is a meaningful clinical problem.

Adult↗
PubMed1989

Structural characterization of the alpha-glycerol-3-phosphate dehydrogenase-encoding gene of Drosophila melanogaster.

In Drosophila, multiple isoforms of alpha-glycerol-3-phosphate dehydrogenase (sn-glycerol-3-phosphate: NAD+ 2-oxidoreductase, EC 1.1.1.8) are produced in a tissue- and stage-specific manner. To understand the underlying molecular basis of these isoforms, we have sequenced a 5.8-kilobase region of the Drosophila genome that contains the entire Gpdh locus. Primer-extension and RNase protection assays show that the gene consists of eight exons and has a single transcription-start point. RNase protection mapping and comparison of the genomic sequence from three different cDNA clones reveal that three protein isoforms of glycerol-3-phosphate dehydrogenase are produced by alternative processing of 3' exons. Two of the isoforms differ from the third by the addition of either three or ten amino acids to their C-terminal ends. Transcripts corresponding to two of the isoforms are expressed during both larval and adult stages, while the third isoform is produced only in adults.

Amino Acid Sequence↗
PubMed1989

Low molecular weight iron from guinea pig reticulocytes isolated by Sephadex G-25 chromatography.

As part of a continuing study of the low MW iron pool, guinea pig reticulocytes were incubated with 59Fe-labeled transferrin, and the reticulocyte hemolysate was chromatographed on Sephadex G-25. 59Fe, in amounts corresponding to that which was in a low MW peak eluting from an Ultrogel column and to that not precipitated by ammonium sulfate, adsorbed to the Sephadex column. The adsorbing 59Fe, on elution from the Sephadex with dilute formic acid, coeluted with phosphate and pentose. When EDTA was added to disrupt the putative iron complex, neither iron nor P adsorbed to the column, supporting the argument that they exist as a compound in the cytosol and adsorb and elute together for that reason. These observations provide additional evidence that P-containing compounds, probably originating as nucleotides, are important components of the low MW iron pool of cells.

Adsorption↗
PubMed1985

Low molecular weight iron in guinea pig reticulocytes.

The low molecular weight iron found in the guinea pig reticulocyte has been partially characterized. On thin layer chromatography it is distinguishable from the iron complexes of a variety of nucleotides, sugars, and amino acids. On paper chromatography it comigrates with a 250-nm absorbing, orcinol-positive material. The eluted count peak contains phosphorus. Approximately 1 microgram of iron is recovered from 1 ml of hemolyzed red cells. Preparation under nitrogen improves recovery of low molecular weight iron, suggesting that the iron is in the ferrous oxidation state.

Animals↗
PubMed1978

Electrostatic complexes of mitomycin C with nucleic acids and polyanions.

Reductively activated mitomycin C exhibits strong, non-covalent electrostatic binding to polyanions such as polyvinylsulfate and polyphosphate. The protonated C-2 amino group generated by the reduction is most likely responsible for this type of interaction. At moderate drug and salt concentrations only covalent binding to nucleic acids is observable. This is shown to be guanine-specific in DNA for the first time, as well as in synthetic polyribo- and polydeoxyribonucleotides at 10--20 times higher binding levels than previously tested. At higher mitomycin C concentration, however, strong non-covalent electrostatic binding to nucleic acids also occurs, resulting in a binding ratio up to 1 mol drug bound per mol mononucleotide, although this non-specific binding is relatively inhibited compared to polyvinylsulfate. Salts also have an inhibitory effect on the non-specific binding to nucleic acids. A series of mitomycin derivatives were compared for their binding and cross-linking abilities using DNA as substrate, with the following results: (a) the presence of a basic nitrogen . funtion at C-2 promotes binding, both covalent and electrostatic, presumably by kinetically facilitating the approach between positively charged nitrogen and DNA. (b) The aziridine ring is the major covalent binding site, indispensable for crosslinking and determines the guanine-specificity of the binding.

Anions↗