Membrane mediated link between ion transport and metabolism in human red cells

When 10^?6 M oubain is added to human red cells that have been incubated without glucose for two hours, there is a significant shift in the ³¹P nuclear magnetic resonances of both phosphate groups of cellular 2,3-diphosphoglycerate, which is not found in control cells incubated with glucose. This means that an effect induced by ouabain on the outside of the red cell membrane is transmitted through the membrane to alter the environment of an intracellular metabolite. Experiments with glycolytic cycle inhibitors have indicated that the intracellular ligand responsible for the resonance shifts is monophosphoglycerate mutase which requires 2,3-diphosphoglycerate as a cofactor for the reaction it catalyzes. To account for this finding a hypothesis is presented that the (Na⁺ + K⁺)-ATPase in human red cells is linked to monophosphoglycerate mutase through the agency of phosphoglycerate kinase. Evidence is presented for the existence of phosphoglycerate kinase/monophosphoglycerate mutase in solution. It is shown that this complex can interact with the cytoplasmic face of (Na⁺ + K⁺)-ATPase at the outside surface of inside out red cell vesicles, and that this interaction is inhibited when 10^?6 M ouabain is contained within the vesicle. Neither monophosphoglycerate mutase nor phosphoglycerate kinase is significantly bound to the inside surface of the intact human red cell, but glyceraldehyde 3-phosphate dehydrogenase is; it is shown that this enzyme also interacts with the cytoplasmic face of the (Na⁺ + K⁺)-ATPase and that the interaction is inhibited by 10^?6 M ouabain.     

Enzymic activity and conformational properties of native and crosslinked glucose oxidase.

Glucose oxidase has been modified by reacting it with glutardialdehyde. The products obtained are water soluble and merely intramolecularly crosslinked; they exhibit high enzymic activity and good stability towards denaturing agents. Comparative circular dichroism studies have been performed with the native and crosslinked enzymes, as well as with the corresponding apoenzymes. The results suggest that the FAD coenzyme is not a gross structural determinant of glucose oxidase.     

Isolation of 3-(2-carboxyethyl)thymine following in vitro reaction of β-propiolactone with calf thymus DNA

3-(2-Carboxyethyl)thymine (3-CET) was synthesized from β-propiolactone (BPL) and dThd5′P at pH 9.0–9.5 via the intermediate 3-(2-carboxyethyl)thymidine-5′-monophosphoric acid (3-CEdThd5′P). 3-CEdThd5′P was converted to 3-CET by hydrolysis in 1.5 N HCl at 100°C for 2 h. The structure of 3-CET was assigned on the basis of UV spectra, electron impact (EI) and isobutane chemical ionization mass spectra and the EI mass spectrum of a trimethylsilyl derivative of 3-CET. BPL was reacted in vitro with calf thymus DNA at pH 7.5. 100 A units of BPL-reacted DNA yielded, following perchloric acid hydrolysis and preparative paper chromatography, 3 A units of 3-CET. Reaction of BPL with the phosphodiester thymidylyl-(3′-5′)thymidine gave 3-(2-carboxyethyl)thymidylyl-(3′-5′)-3-(2-carboxyethyl)thymidine (～3%). Phosphotriester formation was not detected.     

HETEROGENEITY OF HISTAMINE Hi-RECEPTORS: SPECIES VARIATIONS IN [3H]MEPYRAMINE BINDING OF BRAIN MEMBRANES

[³H]Mepyramine binds with high affinity to membranes from brain of human, rat, guinea-pig, rabbit and mouse with drug specificity indicating an association with histamine H₁receptors. Considerable species differences occur in the affinity of [³H]mepyramine, with guinea-pig and human having 34 times greater affinity than rat, mouse or rabbit. The greater affinity of [³H]mepyramine in guinea-pig than in rat is attributable both to faster association and slower dissociation rates in guinea-pig. Species differences in affinity for H₁ receptor sites occur for some antihistamines but not for others. Some tricyclic antidepressant and neuroleptic drugs are extremely potent inhibitors of [³H]mepyramine binding, exceeding in potency any H₁ antihistamines examined. The tricyclic antidepressant doxepin and the neuroleptic clozapine are the most potent of all drugs examined in competing for [³H]mepyramine binding. The regional distribution of specific [³H]mepyramine binding differs considerably in the various species examined.     

Aerometric study of viable fungus spores in an animal care facility.

Studies of airborn fungi were undertaken to evaluate exposure risks for laboratory animals and human handlers which might lead to allergic or invasive disease. Although sporadically high fungus levels were encountered, counts of viable fungus particles were in general low. Recoveries on malt extract agar significantly exceeded those on Sabouraud dextrose agar. The taxa most frequently and abundantly recovered were Penicillium species. Data analyses suggest that 'clean' bedding material may be the principal source of these spores, that cleaning temporarily increases spore levels, and that outdoor airborne fungi contributed little to the indoor air spora identified. Aspergillus fumigatus was infrequently encounted in our samples, and dermatophytes were not recovered.     

Crystallographic data on a complete kappa-type human Bence-Jones protein.

A complete human κ-type Bence—Jones protein (Fin) has been isolated and crystallized. Immunochemical and physicochemical characterization of protein Fin indicates that it is of the κ-chain subgroup, κII, and that it consists of two non-covalently bound intact monomers having a molecular weight of ~23,000 Crystals of Bence—Jones protein Fin obtained from ammonium sulfate solutions have the orthorhombic space group P2₁2₁2₁ with cell dimensions $\text{a = 132.0}\text{A}\text{?}\text{, b = 93.3}\text{A}\text{?}\text{, and}\text{c = 42.3}\text{A}\text{?}$. The asymmetric unit consists of a dimer of molecular weight ~46,000.     

Formation of 15alpha-hydroxyestriol and 15alpha-hydroxyestradiol from C19 15alpha-hydroxylated precursors in human pregnancy.

A mixture of 3H-15alpha-hydroxyandrostenedione and 14C-15alpha-hydroxydehydroisoandrosterone was injected intravenously into two subjects in the third trimester of pregnancy and, in a second study, directly into two fetuses in utero during transfusion for erythroblastosis fetalis. The urine was collected for 4-5 days and steroid conjugates in the urine were hydrolyzed into sulfate and glucosiduronate fractions. From the glucosiduronate fraction 15alpha-hydroxyestriol, 15alpha-hydroxyestradiol, 15alpha-hydroxyandrostenedione and 15alpha-hydroxydehydroisoandrosterone were isolated. No metabolites were identified in the sulfate fraction of the urine. A marked difference was observed in the metabolism of 15alpha-hydroxyandrostenedione and 15alpha-hydroxydehydroisoandrosterone which is dependent on the route of administration of the substrates. Both substrates were converted to 15alpha-hydroxyestriol and 15alpha-hydroxyestradiol, and the 3H/14C ratios and percentage conversions suggest that 15alpha-hydroxyandrostenedione seems to be a better precursor of the urinary 15alpha-hydroxylated estrogens than 15alpha-hydroxydehydroisoandrosterone. The 3H/14C ratios also suggest that 15alpha-hydroxydehydroisoandrosterone was converted to 15alpha-hydroxyestriol via 15alpha-hydroxyandrostenedione, and that the formation of 15alpha-hydroxyestradiol from 15alpha-hydroxydehydroisoandrosterone via 15alpha-hydroxyandrostenedione is a pathway of minor importance. Finally, 15alpha-hydroxydehydroisoandrosterone was recovered from the urine only when the precursors were injected into the maternal circulation. Also, an unknown metabolite containing only 14C was detected in the glucosiduronate fraction of the urine of each subject.     

Catecholamine binding to CNS adrenergic receptors

The properties of ³H-catecholamine binding to α- and β-adrenergic receptors in CNS are reviewed. ³H-epinephrine and ³H-norepinephrine label one class of α-receptors throughout the brain, with high affinities for agonists and some antagonists. Agonist affinities at this site are increased in low temperature conditions but are reduced by guanine nucleotides and monovalent cations. Divalent cations reverse both effects. This α-receptor may be coupled to adenylate cyclase by GTP and/or sodium, and uncoupled by divalent cations. ³H-epinephrine labels β₂, but not β₁, receptors in CNS, especially in bovine cerebellum. The same β-receptor does not show agonist-specific GTP-sensitivity, but does exhibit Na⁺-sensitivity. This receptor appears to be linked to adenylate cyclase, and sodium rather than GTP may be the coupling agent.     

The isolation of peptides by high-performance liquid chromatography using predicted elution positions

This paper describes the derivation and use of predictive retention coefficients for the reversed-phase high-performance liquid chromatography of peptides. The use of predicted elution positions in the isolation of peptides is illustrated by two examples where peptides, whose existence was postulated from cDNA sequence data, have been successfully isolated. The combination of the powerful chromatographic technology and the ability to predict the elution positions of peptides based on their composition provides a very potent method for the isolation of peptides from biological tissues.     

Maximum likelihood estimation of growth yields

This work is concerned with statistical methods to estimate yield and maintenance parameters associated with microbial growth. For a given dilution rate, an experimenter typically measures substrate concentration, oxygen utilization rate, the rate of carbon dioxide evolution, and biomass concentration. These correlated response variables each contain information about the maintenance and yield parameters of interest. A maximum likelihood estimator which combines this correlated information for the yield and maintenance parameters is proposed, evaluated, and tested on literature data. Both point and interval estimators are considered.