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Biomedical subjects

A G Lowe

Publications and source records attributed to A G Lowe.

At least 19 recordsLinked to original sources
PubMed1992

Effects of pressure on glucose transport in human erythrocytes.

The operation of the human red cell glucose transporter has been studied at normal and high hydrostatic pressure to identify the step(s) which involve a volume change. Pressure inhibited zero-trans and equilibrium exchange influx to similar extents, by decreasing the Vmax but not significantly changing the Km. The Bmax and Kd of specific [3H]cytochalasin B binding were unaffected by pressure indicating no change to the number or affinity of functional transporters at pressure. Passive glucose transport was inhibited by pressure in a manner consistent with permeation across the lipid bilayer. These data indicate that there is a major change in volume during the translocation step of the glucose transporter which is rate-limiting for transport.

Biological Transport↗
PubMed1991

Inhibition of glucose transport in human erythrocytes by ubiquinone Q0.

Searches of the protein data bases revealed limited homologies between several regions of the human erythrocyte glucose transporter containing a relative abundance of hydrogen-bonding amino-acid side chains, and proteins of the NADH-ubiquinone oxidoreductase family. This raised the possibility the binding sites for glucose and ubiquinone may be similar in the respective proteins. Experimental studies demonstrated that ubiquinone Q0 does in fact inhibit both glucose entry and glucose exit in human erythrocytes with kinetics consistent with the existence of ubiquinone binding sites at both the exofacial and endofacial sides of the transporter. Glucose transport was also inhibited by the water-soluble tryptophan-inactivating agent, dimethyl(2-hydroxy-5-nitrobenzyl)sulphonium bromide, and this is consistent with the presence of tryptophan residues in two of the exofacial amino-acid sequences proposed as candidates for involvement in glucose binding sites.

Amino Acid Sequence↗
PubMed1987

A single half-turnover of the glucose carrier of the human erythrocyte.

Single half-turnovers of the glucose carrier of the human erythrocyte have been measured by recruiting carriers to the outward-facing conformation by (a) pre-exposing cells to extracellular maltose, or (b) pre-warming cells to 38 degrees C, before addition of D-[14C]glucose at 0 degrees C. Based on these experiments estimates of the number of glucose carriers per red cell range from 124,000 to 190,000.

Blood Glucose↗
PubMed1987

Comparison of the kinetics and thermodynamics of the carrier systems for glucose and leucine in human red blood cells.

Kinetic data for the transport of glucose and leucine in human red blood cells are fitted to the conventional carrier model and the thermodynamics of the two carrier mechanisms are compared. In the absence of the carried molecule both carriers exist mainly in the inward-facing conformation at low temperatures and the outward-facing conformation at physiological or supra-physiological temperatures, this finding reflecting the strongly endothermic process involved in changing from the inward- to outward-facing forms. Reorientations from inward- to outward-conformations also involve substantial increases in entropy for both carriers. In contrast, substrate binding to the glucose carrier involves little change in enthalpy and an increase in entropy, while leucine binding is strongly exothermic and associated with a decrease in entropy. Application of transition state theory to glucose carrier kinetics reveals that the entropy of formation of the transition state of the carrier is much greater than that for the transition state of the carrier-glucose complex.

Carrier Proteins↗
PubMed1986

The kinetics of glucose transport in human red blood cells.

A quenched-flow apparatus and a newly developed automated syringe system have been used to measure initial rates of D-[14C]glucose transport into human red blood cells at temperatures ranging from 0 degrees to 53 degrees C. The Haldane relationship is found to be obeyed satisfactorily at both 0 and 20 degrees C, but Arrhenius plots of maximum D-[14C]glucose transport rates are non-linear under conditions of both equilibrium exchange and zero trans influx. Fitting of the data by non-linear regression to the conventional model for glucose transport gives values at 0 degrees C of 0.726 +/- 0.0498 s-1 and 12.1 +/- 0.98 s-1 for the rate constants governing outward and inward movements of the unloaded carrier molecule and 90.3 +/- 3.47 s-1 and 1113 +/- 494 s-1 for outward and inward movements of the carrier-glucose complex. Activation energies for these four rate constants are respectively 173 +/- 3.10, 127 +/- 4.78, 88.0 +/- 6.17 and 31.7 +/- 5.11 kJ X mol-1. These parameters indicate that at low temperatures the marked asymmetry of the transport mechanism arises mainly from the high proportion of inward-facing carriers and carrier-glucose complexes, and that there is a relatively small difference between the affinities of the carrier for glucose in the inward and outward-facing conformations. At high (physiological) temperatures the carrier is fairly evenly distributed between outward- and inward-facing conformations and the affinity for glucose is about 2.5-times greater outside than inside.

Biological Transport, Active↗
PubMed1985

A quenched-flow technique for the measurement of glucose influx into human red blood cells.

A quenched-flow apparatus is described and applied to measurements of the hydrolysis of 2,4-dinitrophenyl acetate by sodium hydroxide and the entry of D-[U-14C]glucose into human red blood cells at 37 degrees C. Glucose influx into red cells was a saturable process obeying Michaelis-Menten kinetics with a Km for glucose of 6.6 +/- 0.61 mM and a maximum rate for glucose entry under "zero trans" conditions of 20.7 +/- 0.76 mmol (L cell water)-1 s-1. The technique used requires only readily available laboratory equipment and should be easily adaptable to the study of other rapid transport processes.

Biological Transport↗
PubMed1985

Multifit: a flexible non-linear least squares regression program in BASIC.

MULTIFIT, a program in BASIC for implementation on microcomputers, has been developed for non-linear least squares regression fitting of enzyme kinetic, pharmacokinetic and other data to specific models. The program contains a simple procedure for insertion of model equations with up to five parameters (to be fitted) up to 3 independent variables and 1 dependent variable, and can be used to generate a family of programs with pre-set model functions.

Computers↗
PubMed1985

Stimulation of Na+/H+ antiport is an early event in hypertrophy of renal proximal tubular cells.

Renal hypertrophy in vivo is achieved by an increase in protein content per cell and an increase in cell size with minimal hyperplasia. Hypertrophied renal tubular cells remain quiescent and demonstrate an increase in transcellular transport rates. This situation was simulated in vitro by exposing a confluent, quiescent primary culture of rabbit renal proximal tubular cells to either insulin, prostaglandin E1, or hypertonic NaCl for 24 or 48 hr. Protein per cell increased by 20-30% with little or no increase in [3H]thymidine incorporation into DNA. Mean cell volume was also increased in insulin- and hypertonic NaCl-treated but not in prostaglandin E1-treated cells. The lag period required to initiate DNA synthesis by a combination of insulin and hydrocortisone was the same in control and hypertrophied cells, indicating a quiescent state of the latter. Two hours of exposure to the growth stimuli increased amiloride-sensitive Na+ uptake, Na-dependent H+ efflux, and ouabain-sensitive Rb+ uptake, indicating that stimulation of Na+/H+ antiport (exchange) occurs as an early event in their action. Hypertrophied cells continued to demonstrate enhanced Na+/H+ antiport after the growth stimuli were removed for 3 hr, by which time their acute effects are reversed.

Alprostadil↗
PubMed1978

Chloride/bicarbonate exchange in human erythrocytes.

1. The exchange of chloride and bicarbonate across the human erythrocyte membrane has been followed by measuring the changes in extracellular pH which occur when chloride-rich erythrocytes are added to chloride-free media containing varying concentrations of bicarbonate and carbonic anhydrase. 2. The dependence of the rate of chloride/bicarbonate exchange on the extracellular concentration of bicarbonate was consistent with the existence of a saturable membrane anion transporter exhibiting Michaelis--Menten kinetics. In a medium containing sodium gluconate buffered to pH 7.0 with imidazole--malate the Km for bicarbonate activation of transport was 0.39 (+/- 0.03) mM and the Vmax was 2033 (+/- 80 m-mole anions exchanged/3 X 10(13) cells. min, at 10 degrees C. 3. Chloride/bicarbonate exchange was temperature-dependent with an Arrhenius activation energy of 19.4 kcal/mole in the temperature range 2--10 degrees C. 4. Exchange of intracellular chloride for extracellular bicarbonate was inhibited by the presence of extracellular halides. Inhibition by chloride, bromide and fluoride was competitive and the affinity of the transport system decreased in the order HCO-3 greater than Cl- greater than Br- greater than F-. The kinetics of inhibition by iodide were complex, but inhibitory effects of low concentrations of iodide were less than those of chloride and bromide.

Bicarbonates↗
PubMed1977

The pre-steady-state hydrolysis of ATP by porcine brain (Na+ + K+)-dependent ATPase.

The hydrolysis of [gamma-32P]ATP by porcine brain (Na+ + K+)-stimulated ATP phosphohydrolase (EC 3.6.1.3) has been studied at 28 degree C in a rapid mixing quenched-flow apparatus. An "early burst" in the release of Pi from ATP has been observed when the enzyme is mixed with ATP, Na+ and a relatively high concentration of K+ (10 mM) but the burst is less pronounced with 0.5 mM K+. This "early burst" of Pi release is suppressed when the enzyme is pre-mixed with 10 mM K+ or 20% (v/v) dimethylsulphoxide before mixing with ATP and Na+, and premixing of enzyme with Na+ antagonizes this effect of dimethylsulphoxide. The results have been analysed by a non-linear least squares regression treatment and are consistent with a mechanism involving three steps, one of which may be a relatively slow change in enzyme conformation following release of Pi from its covalent linkage with the enzyme, in addition to formation of the enzyme-substrate complex. Rate constants (and S.E.) for these steps have been calculated and the roles of phospho-enzyme and other intermediates in the reaction mechanism of the transport ATPase are dicussed.

Adenosine Triphosphatases↗