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Biomedical subjects

W W Kay

Publications and source records attributed to W W Kay.

At least 91 records · Page 5Linked to original sources
PubMed1981

Inorganic cation transport and the effects on C4 dicarboxylate transport in Bacillus subtilis.

The complex interrelationships between the transport of inorganic cations and C4 dicarboxylate were examined using mutants defective in potassium transport and retention, divalent cation transport, or phosphate transport. The potassium transport system, studied using 86Rb+ as a K+ analogue, kinetically appeared as a single system (Km 200 microM for Rb+, Ki 50 microM for K+), the activity of which was only slightly reduced in K+ retention mutants. Divalent cation transport, studied using 54Mn2+, 60Co2+, and 45Ca2+, was more complex being represented by at least two systems, one with a high affinity for Mn2+ (Km 2.5 microM) and a more general one of low affinity (Km 1.3-10 mM) for Mg2+, Mn2+, Ca/2+, and Co2+. Divalent cation transport was repressed by Mg2+, derepressed in K+ retention mutants, and defective in Co2+-resistant mutants. Phosphate was required for both divalent cation and succinate transport, and phosphate transport mutants (arsenate resistant) were found to be defective in both divalent cation and succinate transport. Divalent cations, especially Mg2+ and Co2+, decreased Km for succinate transport approximately 20-fold over that achieved with K+; neither cation was required stoichiometrically for succinate transport. The loss of divalent cation transport in cobalt-resistant mutants has been correlated with the loss of a 55,000 molecular weight membrane protein. Similarly, the loss of phosphate transport in arsenate-resistant mutants has been correlated with the loss of a 35,000 molecular weight membrane component.

Bacillus subtilis↗
PubMed1980

Citrate transport in Salmonella typhimurium: studies with 2-fluoro-L-erythro-citrate as a substrate.

The citrate analogue, 2-fluoro-L-erythro-[3,4,5,6-(14)C]citrate was synthesized as a probe for the citrate transport system of Salmonella typhimurium. This analogue was actively transported by an inducible energy-dependent transport system with high affinity for fluorocitrate (Km = 3.3 microM), and this transport system was inhibited competitively by citrate and isocitrate. Fluorocitrate was shown to be a competitive inhibitor of the citrate-binding protein (C protein) of this organism (Ki = 4-5 microM). Analogue resistant mutants were simultaneously defective in fluorocitrate transport as well as the C protein and the affected allele, tctC, was located at 59 units on the S. typhimurium chromosome map. These tctC mutants were shown to be specifically defective in K+-dependent fluorocitrate transport but still retained another system capable of transporting fluorocitrate in the presence of both Na+ and K+.

Binding, Competitive↗
PubMed1979

Purification and properties of a citrate-binding transport component, the C protein of Salmonella typhimurium.

Salmonella typhimurium was shown to contain a citrate-binding protein (C protein) which was purified to homogeneity from the periplasmic fraction released by cold osmotic shock. The protein is dimeric, has an apparent molecular weight of 28 000 and an isoelectric point of 6.1. Sodium ions were required for optimum substrate binding, however, the divalent cations Zn2+, Mg2+, and Co2+ were inhibitory. The C protein was relatively stable but sensitive to various detergents and chaotropic agents. Approximately one citrate molecule was bound per molecule of protein and citrate binding (Kd = 1-2.6 microM) was strongly competitively inhibited by DL-isocitrate and DL-fluorocitrate but not by other carboxylates. Neither succinate, glutamate, nor acetate were bound to the C protein. No apparent enzyme activity was associated with this protein. A concomitant reduction in the level of binding protein and in citrate transport activity occurred in osmotically shocked cells as well as with L-malate- or succinate-grown cells. Fluorocitrate-resistant mutants were simultaneously defective in citrate transport, citrate binding, and production of cross-reacting material. One transport-defective mutant did produce citrate binding protein.

Biological Transport, Active↗
PubMed1977

Binding of collagen by canine blood platelets.

The binding of 125I-collagen (tropocollagen) by canine blood platelets occurred in a concentration-dependent manner but no saturation effect could be observed. The binding of collagen was not entirely specific for platelets since various eucaryotic and procaryotic cells quantitatively bound collagen as well or better. The temporal response to added collagen appeared to be binding, 3H-serotonin release, and finally platelet aggregation. Non-polymerizing salt-soluble tropocollagen was bound as well as acid-soluble tropocollagen, however neither 3H-serotonin release nor platelet aggregation occurred. Furthermore, the binding activity was not destroyed by treatment with collagenase, galactose oxidase and glucose oxidase, nor by periodate oxidation. Platelet aggregation closely paralleled acid soluble collagen polymerization and both events were equally inhibited by arginine; however, arginine did not interfere with collagen binding. Scanning electron microscopy revealed an unusual morphological platelet response to collagen and platelets appeared to be nucleation sites for collagen polymerization.

Animals↗
PubMed1977

Inhibition and enhancement of platelet aggregation by collagen derivatives.

Rat tail tendon tropocollagen rapidly caused the aggregation of blood platelets coincident with collagen multimerization. Several long-chain alkyl derivatives of collagen, however, were completely unable to induce platelet aggregation even at unusually high concentrations. These collagen derivatives varied in their polymerization properties , and the polymerization of tropocollagen derivatives was shown not necessarily to be a prerequisite for platelet aggregation. These collagen derivatives may enhance the antithrombogenic properties of collagenbased prosthese. A short-chain derivative, collagen-glycine ethyl ester, profoundly enhanced platelet aggregation in the absence of any measurable polymerization. This derivative may be useful as a growth-promoting burn dressing.

Animals↗
PubMed1975

Succinate transport in Bacillus subtilis. Dependence on inorganic anions.

Cations were generally ineffective in stimulating succinate transport in a succinate dehydrogenase mutant of Bacillus subtilis unless accompanied by polyvalent anions; phosphate and sulfate being particularly active. The Km values for the phosphate or sulfate requirement were approx. 3 mM. Biphasic kinetics were characteristic of both the succinate (Km values 0.1 and 1 mM), and inorganic phosphate (Km values 0.1 and 3 mM) transport system(s). The phosphate transport system(s) was repressed by high inorganic phosphate and a coordinate increase in the transport of phosphate, arsenate, and phosphate-stimulated succinate transport accompanied growth in low phosphate media. A class of arsenate resistant mutants were simultaneously defective in the transport of arsenate, phosphate and succinate when cells were repressed for phosphate transport, however, the transport of these ions was regained in these mutants when grown in low phosphate media. Organic phosphate esters did not stimulate succinate transport in arsenate resistant mutants but were effective after growth in low phosphate media. Growth under phosphate limitation permitted the simultaneous regain of both phosphate and sulfate dependent succinate transport activities whereas sulfate limitation alone was ineffective. Succinate was not transported by an anion exchange diffusion mechanism since phosphate efflux was low or absent during succinate transport. The transport of C4-dicarboxylates in B. subtilis is strongly stimulated by intracellular polyvalent anions. The absence of an anion permeability mechanism precludes succinate transport but partial escape from this restriction is mediated by the derepression of a phosphate transport system.

Arsenates↗
PubMed1975

The fate of autologous fascia lata grafts in dogs.

The fate of tricuspid fascia lata valves and intracardiac grafts was studied morphologically, histologically, and biochemically in dogs. Severe degenerative changes occurring by 21 days were largely correlated with graft thickening due to gross endothelialization. A primary event in graft alteration was an initial loss of the indigenous fibroblast population as detected both microscopically and by autoradiography after pulse labeling with 14C-Nethylmaleimide. Using different isotopic labeling procedures, both the in vivo and in vitro graft degradation patterns demonstrated that a second early eft degradation was correlated in vivo with endothelialization and collagenase secretion by invasive cells.

Animals↗
PubMed1975

Biochemical modifications of fascia lata for autologous valve replacement.

The biochemistry of canine fascia lata was studied to assess the feasibility of tissue modification with a variety of cross-linking reagents and growth inhibitors in order to provide a more suitable material for heart valves. Chemical determinants for maximal reactivity were consequently established for several growth inhibitors, of which the hydrophobic ligands difluorodinitrobenzene, octadecylamine, and oleylamine were the most effective in preventing invasive cellular growth. This enhanced the longevity of atrial grafts in dogs as judged from morphological and histological examination of excised grafts.

Amino Acids↗
PubMed1975

Salmonella typhimurium HfrA, a mutant in which adenosine triphosphate can drive amino acid transport but not energy-dependent nicotinamide nucleotide transhydrogenation.

In contrast with wild-type Salmonella typhimurium LT2, strain HfrA did not have ATP-driven energy-dependent transhydrogenase activity, although ATP-dependent quenching of atebrin fluorescence was normal. Respiration-dependent and energy-independent transhydrogenase, and Ca2+-activated ATPase (adenosine triphosphatase) activities were similar in both strains. Purified ATPases from the two strains had similar specific activities, similar subunit polypeptides, and were equally effective in restoring energy-dependent transhydrogenase activities to membrane particles of strain LT2 from which the ATPase had been stripped. The purified ATPases from both strains could restore respiration-dependent but not ATP-dependent transhydrogenation to stripped particles of strain HfrA. Both strains grew aerobically equally well on salts media containing glucose, malate, succinate, citrate, acetate, pyruvate, fumarate, lactate or aspartate as substrates. Growth on glucose under anaerobic conditions was similar. Strains LT2 and HfrA were equally effective in the accumulation under both aerobic and anaerobic conditions of the amino acids proline, phenylalanine, histidine, lysine, isoleucine and aspartic acid. Inhibition of amino acid accumulation by KCN and dicyclohexylcarbodi-imide occurred to the same extent in both strains. The complete inhibition by dicyclohexylcarbodi-imide of amino acid uptake under anaerobic conditions suggested that ATP could drive amino acid uptake in both strains. The ability of strain HfrA to carry out ATP-dependent transport or quenching of atebrin fluorescence but not ATP-dependent transhydrogenation is different from the wild-type strain and from any previously described energy-coupling mutant. It is difficult to reconcile the properties of this mutant with the chemiosmotic hypothesis.

Adenosine Triphosphatases↗
PubMed1975

Regulation of C4-dicarboxylic acid transport in Bacillus subtilis.

Various mutant strains of Bacillus subtilis were used to study the induction and regulation of the transport of tricarboxylic acid cycle C4-dicarboxylates. L-Malate was the only physiological inducer and bromosuccinate was a gratuitous inducer of dicarboxylic acid transport in a succinic dehydrogenase deficient mutant. Several mutants were isolated with alterations in the ability to transport dicarboxylic acids. One of these (WK6) was defective in the ability to take up succinate when grown on glutamate minimal medium, whereas another (WK1) was inducible to high levels by extremely low levels of L-malate. Alpha-Ketoglutarate dehydrogenase mutants were unable to take up dicarboxylates because of repression of transport by glutamate and (or) alpha-ketoglutarate. A mutation which resulted in increased levels of alpha-ketoglutarate dehydrogenase partially overcame this inhibition. Glutamate did not prevent the induction of dicarboxylic acid transport by L-malate in succinic dehydrogenase mutants but was markedly inhibitory in alpha-ketoglutarate dehydrogenase mutants.

Aspartic Acid↗
PubMed1973

Properties of an inducible C 4 -dicarboxylic acid transport system in Bacillus subtilis.

The transport of the tricarboxylic acid cycle C(4)-dicarboxylic acids was studied in both the wild-type strain and tricarboxylic acid cycle mutants of Bacillus subtilis. Active transport of malate, fumarate, and succinate was found to be inducible by these dicarboxylic acids or by precursors to them, whereas glucose or closely related metabolites catabolite-repressed their uptake. l-Malate was found to be the best dicarboxylic acid transport inducer in succinic dehydrogenase, fumarase, and malic dehydrogenase mutants. Succinate and fumarate are accumulated over 100-fold in succinic dehydrogenase and fumarase mutants, respectively, whereas mutants lacking malate dehydrogenase were unable to accumulate significant quantities of the C(4)-dicarboxylic acids. The stereospecificity of this transport system was studied from a comparison of the rates of competitive inhibition of both succinate uptake and efflux in a succinate dehydrogenase mutant by utilizing thirty dicarboxylic acid analogues. The system was specific for the C(4)-dicarboxylic acids of the tricarboxylic acid cycle, neither citrate nor alpha-ketoglutarate were effective competitive inhibitors. Of a wide variety of metabolic inhibitors tested, inhibiors of oxidative phosphorylation and of the formation of proton gradients were the most potent inhibitors of transport. From the kinetics of dicarboxylic acid transport (K(m) approximately 10(-4) M for succinate or fumarate in succinic acid dehydrogenase and fumarase mutants) and from the competitive inhibition studies, it was concluded that an inducible dicarboxylic acid transport system mediates the entry of malate, fumarate, or succinate into B. subtilis. Mutants devoid of alpha-ketoglutarate dehydrogenase were shown to accumulate both alpha-ketoglutarate and glutamate, and these metabolites subsequently inhibited the transport of all the C(4)-dicarboxylic acids, suggesting a regulatory role.

Aspartic Acid↗