Denitrifying Pseudomonas aeruginosa: some parameters of growth and active transport.
(1/531)
Optimal cell yield of Pseudomonas aeruginosa grown under denitrifying conditions was obtained with 100 mM nitrate as the terminal electron acceptor, irrespective of the medium used. Nitrite as the terminal electron acceptor supported poor denitrifying growth when concentrations of less than 15 mM, but not higher, were used, apparently owing to toxicity exerted by nitrite. Nitrite accumulated in the medium during early exponential phase when nitrate was the terminal electron acceptor and then decreased to extinction before midexponential phase. The maximal rate of glucose and gluconate transport was supported by 1 mM nitrate or nitrite as the terminal electron acceptor under anaerobic conditions. The transport rate was greater with nitrate than with nitrite as the terminal electron acceptor, but the greatest transport rate was observed under aerobic conditions with oxygen as the terminal electron acceptor. When P. aeruginosa was inoculated into a denitrifying environment, nitrate reductase was detected after 3 h of incubation, nitrite reductase was detected after another 4 h of incubation, and maximal nitrate and nitrite reductase activities peaked together during midexponential phase. The latter coincided with maximal glucose transport activity. (+info)
Intrarenal site of action of calcium on renin secretion in dogs.
(2/531)
We studied the effects of intrarenal calcium infusion on renin secretion in sodium-depleted dogs in an attempt to elucidate the major site of calcium-induced inhibition of renin release. Both calcium chloride and calcium gluconate reduced renal blood flow and renin secretion while renal perfusion pressure was unchanged. These data indicate that calcium inhibition of renin secretion did not occur primarily at the renal vascular receptor; decreased renal blood flow is usually associated with increased renin secretion. Calcium chloride infusion increased urinary chloride excretion without affecting sodium excretion, and calcium gluconate failed to increase either sodium or chloride excretion. Also, the filtered loads of sodium and chloride were unchanged during the calcium infusions. These results give no indication that calcium inhibited renin secretion by increasing the sodium or chloride load at the macula densa. The effects of intrarenal calcium infusion on renin release were also assessed in dogs with a nonfiltering kidney in which renal tubular mechanisms could not influence renin secretion. The observation that calcium still suppressed renin release in these dogs provides additional evidence that the the major effect of calcium involved nontubular mechanisms. Thus, it appears likely that calcium acted directly on the juxtaglomerular cells to inhibit renin secretion. (+info)
A kinetic study of ribulose bisphosphate carboxylase from the photosynthetic bacterium Rhodospirillum rubrum.
(3/531)
The activation kinetics of purified Rhodospirillum rubrum ribulose bisphosphate carboxylase were analysed. The equilibrium constant for activation by CO(2) was 600 micron and that for activation by Mg2+ was 90 micron, and the second-order activation constant for the reaction of CO(2) with inactive enzyme (k+1) was 0.25 X 10(-3)min-1 . micron-1. The latter value was considerably lower than the k+1 for higher-plant enzyme (7 X 10(-3)-10 X 10(-3)min-1 . micron-1). 6-Phosphogluconate had little effect on the active enzyme, and increased the extent of activation of inactive enzyme. Ribulose bisphosphate also increased the extent of activation and did not inhibit the rate of activation. This effect might have been mediated through a reaction product, 2-phosphoglycolic acid, which also stimulated the extent of activation of the enzyme. The active enzyme had a Km (CO2) of 300 micron-CO2, a Km (ribulose bisphosphate) of 11--18 micron-ribulose bisphosphate and a Vmax. of up to 3 mumol/min per mg of protein. These data are discussed in relation to the proposed model for activation and catalysis of ribulose bisphosphate carboxylase. (+info)
Relationship between catabolism of glycerol and metabolism of hexosephosphate derivatives by Pseudomonas aeruginosa.
(4/531)
The relationship between catabolism of glycerol and metabolism of hexosephosphate derivatives in Pseudomonas aeruginosa was studied by comparing the growth on glycerol and enzymatic constitution of strain PAO with these characteristics of glucose-catabolic mutants and revertants. Growth of strain PAO on glycerol induced a catabolic oxidized nicotinamide adenine dinucleotide-linked glyceraldehyde-phosphate dehydrogenase and seven glucose-catabolic enzymes. The results indicated that these enzymes were induced by a six-carbon metabolite of glucose. All strains possessed a constitutive anabolic Embden-Meyerhof-Parnas pathway allowing limited conversion of glycerol-derived triosephosphate to hexosephosphate derivatives, which was consistent with induction of these enzymes by glycerol. Phosphogluconate dehydratase-deficient mutants grew on glycerol. However, mutants lacking both phosphogluconate dehydrogenase and phosphogluconate dehydratase were unable to grow on glycerol, although these strains possessed all of the enzymes needed for degradation of glycerol. These mutants apparently were inhibited by hexosephosphate derivatives, which originated from glycerol-derived triosephosphate and could not be dissimilated. This conclusion was supported by the fact that revertants regaining only a limited capacity to degrade 6-phosphogluconate were glycerol positive but remained glucose negative. (+info)
Transport characteristics of the apical anion exchanger of rabbit cortical collecting duct beta-cells.
(5/531)
To functionally characterize transport properties of the apical anion exchanger of rabbit beta-intercalated cells, the mean change in anion exchange activity, dpHi/dt (where pHi is intracellular pH), was measured in response to lumen Cl- replacement with gluconate in perfused cortical collecting ducts (CCDs). beta-Cell apical anion exchange was not affected by 15-min exposure to 0.2 mM lumen DIDS in the presence of 115 mM Cl-. In contrast, apical anion exchange was significantly inhibited by 0.1 mM lumen DIDS in the absence of Cl-. beta-Cell apical anion exchange was unchanged by 15 mM maleic anhydride, 10 mM phenylglyoxal, 0.2 mM niflumic acid, 1 mM edecrin, 1 mM furosemide, 1 mM probenecid, or 0.1 mM diphenylamine-2-carboxylate. However, beta-cell apical anion exchange was inhibited by alpha-cyano-4-hydroxycinnamic acid, with an IC50 of 2.4 mM. Substitution of either sulfate or gluconate for lumen Cl- resulted in a similar rate of alkalinization. Conversely, pHi was unchanged by substitution of sulfate for lumen gluconate, confirming the lack of transport of sulfate on the beta-cell apical anion exchanger. Taken together, the results demonstrate a distinct "fingerprint" of the rabbit CCD beta-cell apical anion exchanger that is unlike that of other known anion exchangers. (+info)
Common components of patch-clamp internal recording solutions can significantly affect protein kinase A activity.
(6/531)
Common components of whole-cell internal recording solutions were tested both in vitro and in patch-clamp experiments for their effects on the activity of cAMP-dependent protein kinase. Potassium fluoride (KF), 440 mM trimethylamine chloride and exclusion of bovine serum albumin (BSA) decreased the activity of the enzyme, while ethylene glycol-bis (beta-aminoethyl ether) N,N,N',N'-tetraacetic acid (EGTA) and the potassium salts of aspartate, gluconate, methylsulfate and monobasic phosphate increased its activity. Addition of KF to the internal solution produced a hyperpolarizing shift in the V1/2 of Ih channel activation, consistent with the KF-induced reduction of protein kinase A activity. Therefore, consideration of the composition of internal solutions is warranted when studying channel physiology by patch-clamp techniques. (+info)
Inhibition of P-glycoprotein-mediated transport by a hydrophobic contaminant in commercial gluconate salts.
(7/531)
The substitution of gluconate for Cl- is commonly used to characterize Cl- transport or Cl--dependent transport mechanisms. We evaluated the effects of substituting gluconate for Cl- on the transport of the P-glycoprotein substrate rhodamine 123 (R123). The replacement of Ringer solution containing Cl- (Cl--Ringer) with gluconate-Ringer inhibited R123 efflux, whereas the replacement of Cl- by other anions (sulfate or cyclamate) had no effect. The inhibition of R123 efflux by gluconate-Ringer was absent after chloroform extraction of the sodium gluconate salt. The readdition of the sodium gluconate-chloroform extract to the extracted gluconate-Ringer or to cyclamate-Ringer inhibited R123 efflux, whereas its addition to Cl--Ringer had no effect. These observations indicate that the inhibition of P-glycoprotein-mediated R123 transport by gluconate is due to one or more chloroform-soluble contaminants and that the inhibition is absent in the presence of Cl-. The results are consistent with the fact that P-glycoprotein substrates are hydrophobic. Care should be taken when replacing ions to evaluate membrane transport mechanisms because highly pure commercial preparations may still contain potent contaminants that affect transport. (+info)
Evidence for chloride ions as intracellular messenger substances in astrocytes.
(8/531)
Cultured rat hippocampal astrocytes were used to investigate the mechanism underlying the suppression of Ba2+-sensitive K+ currents by GABAA receptor activation. Muscimol application had two effects on whole cell currents: opening of the well-known Cl- channel of the GABAA receptor and a secondary longer-lasting blockade of outward K+ currents displaying both peak and plateau phases. This blockade was independent of both Na+ (inside and outside) and ATP in the pipette. It also seemed to be independent of muscimol binding to the receptor because picrotoxin application showed no effect on the K+ conductance. The effect is blocked when anion efflux is prevented by replacing Cl- with gluconate (both inside and out) and is enhanced with more permeant anions such as Br- and I-. Moreover, the effect is reproduced in the absence of muscimol by promoting Cl- efflux via lowering of extracellular Cl- levels. These results, along with the requirement for Cl- efflux in muscimol experiments, show a strong dependency of the secondary blockade on Cl- efflux through the Cl- channel of the GABAA receptor. We therefore conclude that changes in the intracellular Cl- concentration alter the outward K+ conductances of astrocytes. Such a Cl--mediated modulation of an astrocytic K+ conductance will have important consequences for the progression of spreading depression through brain tissue and for astrocytic swelling in pathological situations. (+info)