Correlation of microbial mass with ATP and DNA concentrations in acidogenesis of whey permeate

In this paper, we examine variations in the contents of ATP and DNA per unit microbial mass in an acidogenesis of whey permeate. We also introduce a novel approach to estimate microbial mass by measuring ATP and DNA when the ratios of ATP and DNA to microbial mass vary. Acidogenic experiments were performed at 35°C and pH 6.0 in batch mode. The amounts of ATP and DNA per unit microbial mass were not consistent during the incubation except during the post-decay phase. Especially within the exponential phase, each showed a 10-fold difference between maximal and minimal values. In this case, the conventional method which converts ATP or DNA concentration into microbial mass using a fixed conversion factor can give inaccurate results. While the constant ratios of 0.74 mg ATP/g VSS and 1.96 mg DNA/g VSS were determined for the post-decay phase, the ATP and DNA concentrations showed strong linear relationships with the microbial mass (r ² = 0.99) within the ranges of 0.039–1.078 mg ATP/l and 0.075–2.080 mg DNA/l, respectively. The linear regression equations are as follows: (1) microbial mass concentration (mg/l) = 478.5 × ATP concentration (mg/l) + 293.5, (2) microbial mass concentration (mg/l) = 257.2 × DNA concentration (mg/l) + 250.4. Therefore, changes in the mass of the acidogenic population should be monitored by the combined use of the regression equations obtained in the exponential phase and the constant ratios determined in the post-decay phase. This procedure should be widely applicable to the acidogenesis of dairy processing wastewaters, especially of a highly suspended organic wastewater such as whey.     

ATP interaction with the open state of the K(ATP) channel

The mechanism of ATP-sensitive potassium (K(ATP)) channel closure by ATP is unclear, and various kinetic models in which ATP binds to open or to closed states have previously been presented. Effects of phosphatidylinositol bisphosphate (PIP2) and multiple Kir6.2 mutations on ATP inhibition and open probability in the absence of ATP are explainable in kinetic models where ATP stabilizes a closed state and interaction with an open state is not required. Evidence that ATP can in fact interact with the open state of the channel is presented here. The mutant Kir6.2[L164C] is very sensitive to Cd2+ block, but very insensitive to ATP, with no significant inhibition in 1 mM ATP. However, 1 mM ATP fully protects the channel from Cd2+ block. Allosteric kinetic models in which the channel can be in either open or closed states with or without ATP bound are considered. Such models predict a pedestal in the ATP inhibition, i.e., a maximal amount of inhibition at saturating ATP concentrations. This pedestal is predicted to occur at >50 mM ATP in the L164C mutant, but at >1 mM in the double mutant L164C/R176A. As predicted, ATP inhibits Kir6.2[L164C/R176A] to a maximum of approximately 40%, with a clear plateau beyond 2 mM. These results indicate that ATP acts as an allosteric ligand, interacting with both open and closed states of the channel.     

The effects of heavy metals on microbial biomass in sediments of Palestine Lake

Effects of metal contamination on microbial biomass in sediment samples from three areas in Palestine Lake (one area highly polluted with chromium, cadmium and zinc) were determined. Adenosine triphosphate (ATP) concentrations, determined by the luciferin-luciferase bioluminescent technique, and microbial colony numbers on pour plates were used as biomass indicators. Plate counts showed a significant (P < 0.01) site effect with the highly contaminated area having an order of magnitude lower microbial population than the control area. ATP concentrations also indicated lower microbial biomass in contaminated sediments. The metal concentrations of the most contaminated area averaged 17,840 µg Zn/g, 4380 µg Cr/g and 585 µg Cd/g based on dry weight of sediments. A suppression of organic decomposition was evident in the impacted area; high metal levels and resultant low microbial biomass may have been causative.     

Effect of ATP concentration on CFTR Cl- channels: a kinetic analysis of channel regulation.

Phosphorylated cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channels require nucleoside triphosphates, such as ATP, to open. As the concentration of intracellular ATP increases, the probability of the channel being open (Po) increases. To better understand how ATP regulates the channel, we studied excised inside-out membrane patches that contained single, phosphorylated CFTR Cl- channels and examined the kinetics of gating at different concentrations of ATP. As the ATP concentration increased from 0.1 to 3 mM the mean closed time decreased, but mean open time did not change. Analysis of the data using histograms of open- and closed-state durations, the maximum likelihood method, and the log-likelihood ratio test suggested that channel behavior could be described by a model containing one open and two closed states (C1<==>C2<==>O). ATP regulated phosphorylated channels at the transition between the closed states C1 and C2: as the concentration of ATP increased, the rate of transition from C1 to C2 (C1-->C2) increased. In contrast, transitions from C2 to C1 and between C2 and the open state (O) were not significantly altered by ATP. Addition of ADP in the presence of ATP decreased the transition rate from C1 to C2 without affecting other transition rates. These data suggest that ATP regulates CFTR Cl- channels through an interaction that increases the rate of transition from the closed state to a bursting state in which the channel flickers back and forth between an open and a closed state (C2). This transition may reflect ATP binding or perhaps a step subsequent to binding.     

Biochemical indication of microbial mass changes using ATP and DNA measurement in biological treatment of thiocyanate

This study was designed to monitor changes in the levels of adenosine 5'-triphosphate (ATP) and deoxyribonucleic acid (DNA) per unit of microbial mass during the autotrophic biodegradation of thiocyanate (SCN(-)). An artificial medium containing trace minerals and 500 mg SCN(-)/L was used as a substrate for bacterial growth. An SCN(-)-degrading bioreactor with a working volume of 6 L, equipped with temperature, pH, and dissolved oxygen controls, was operated in batch mode. During the exponential phase of SCN(-) biodegradation, the ratios of ATP and DNA to microbial dry weight varied from 0.6 to 1.1 mug ATP/mg of volatile suspended solid (VSS), and from 3.5 to 8.8 mug DNA/mg of VSS, respectively. The ATP and DNA concentrations correlated linearly with microbial mass (r (2) > 0.9) within the exponential phase. The linear regression equations were as follows: (1) microbial mass concentration (mg/L) = 0.663 x ATP concentration (mug/L) + 11.1 and (2) microbial concentration (mg/L) = 0.081 x DNA concentration (mug/L) + 10.9. The applicable ranges were 6.8 to 47.4 mug/L for ATP concentration and 41.5 to 395 mug/L for DNA concentration, respectively.     

Dual effect of adenosine triphosphate on the apical small conductance K+ channel of the rat cortical collecting duct

We used the patch-clamp technique to study the effects of ATP on the small-conductance potassium channel in the apical membrane of rat cortical collecting duct (CCD). This channel has a high open probability (0.96) in the cell-attached mode but activity frequently disappeared progressively within 1-10 min after channel excision (channel "run-down"). Two effects of ATP were observed. Using inside-out patches, low concentrations of ATP (0.05-0.1 mM) restored channel activity in the presence of cAMP-dependent protein kinase A (PKA). In contrast, high concentrations (1 mM) of adenosine triphosphate (ATP) reduced the open probability (Po) of the channel in inside-out patches from 0.96 to 0. 1.2 mM adenosine diphosphate (ADP) also blocked channel activity completely, but 2 mM adenosine 5'-[beta,gamma-imido]triphosphate (AMP-PNP), a nonhydrolyzable ATP analogue, reduced Po only from 0.96 to 0.87. The half-maximal inhibition (Ki) of ATP and ADP was 0.5 and 0.6 mM, respectively, and the Hill coefficient of both ATP and ADP was close to 3. Addition of 0.2 or 0.4 mM ADP shifted the Ki of ATP to 1.0 and 2.0 mM, respectively. ADP did not alter the Hill coefficient. Reduction of the bath pH from 7.4 to 7.2 reduced the Ki of ATP to 0.3 mM. In contrast, a decrease of the free Mg2+ concentration from 1.6 mM to 20 microM increased the Ki of ATP to 1.6 mM without changing the Hill coefficient; ADP was still able to relieve the ATP-induced inhibition of channel activity over this low range of free Mg2+ concentrations. The blocking effect of ATP on channel activity in inside-out patches could be attenuated by adding exogenous PKA catalytic subunit to the bath. The dual effects of ATP on the potassium channel can be explained by assuming that (a) ATP is a substrate for PKA that phosphorylates the potassium channel to maintain normal function. (b) High concentrations of ATP inhibit the channel activity; we propose that the ATP-induced blockade results from inhibition of PKA-induced channel phosphorylation.     

Analysis of Plankton Dynamics in a Southwestern U.S.A. Reservoir Using ATP Assays

Plankton biomass in size segregated (i.e. 0.45–10 μm, 10–64 μm, 64–165 μm, and > 165 μm) samples was measured using ATP assays for two years in H. H. Moss Reservoir. The presence of variable numbers of microcrustacean zooplankton in the > 165 μ size class introduced significant, and perhaps inaccurate, variance into temporal dynamics of total ATP concentrations. As many as six physiochemical parameters, measured during 1975–76, were required to produce a significant multiple regression against ATP concentrations; a significant correlation between total ATP measured in the photic zone and surface temperature was observed. A large standing crop of nannoplankton (i.e. 0.45–10 μm) existed in the anaerobic and aphotic hypolimnion during summer stratification both years. This suggested that microbial heterotrophy was an especially important primary trophic event in the reservoir. Particulate (POC) and dissolved (DOC) organic carbon were measured in 1976–77. Organic carbon (POC + DOC) regressed significantly against ATP concentration in the 0.45–10 μm size class and microbial biomass (i.e. 0.45–165 μm) averaged 32 per cent of POC. When biomass was monitored over 24-hour periods in the photic zone, significant changes were documented in various size classes, especially nannoplankton; but total microbial biomass remained relatively constant.     

A rapid method for measuring ATP + ADP + AMP in marine sediment

In this report, I describe a method for rapid measurement of total adenylate (ATP + ADP + AMP) in marine sediment samples for estimating microbial biomass. A simple ‘boil and dilute’ method is described here, whereby adding boiled MilliQ water to sediments increases the detection limit for ATP + ADP + AMP up to 100-fold. The lowered detection limit of this method enabled the detection ATP + ADP + AMP in relatively low-biomass sub-seafloor sediment cores with 10⁴ 16S rRNA gene copies per gram. Concentrations of ATP + ADP + AMP correlated with 16S rRNA gene concentrations from bacteria and archaea across six different sites that range in water depth from 1 to 6000 m indicating that the ATP + ADP + AMP method can be used as an additional biomass proxy. In deep sea microbial communities, the ratio of ATP + ADP + AMP concentrations to 16S rRNA genes >1 m below seafloor was significantly lower compared to communities in the upper 30 cm of sediment, which may be due to reduced cell sizes and or lower ATP + ADP + AMP concentrations per cell in the deep sea sub-seafloor biosphere. The boil and dilute method for ATP + ADP + AMP is demonstrated here to have a detection limit sufficient for measuring low biomass communities from deep sea sub-seafloor cores. The method can be applied to frozen samples, enabling measurements of ATP + ADP + AMP from frozen sediment cores stored in core repositories from past and future international drilling campaigns.     

ATP and Ergosterol as Indicators of Fungal Biomass during Leaf Decomposition in Streams: a Comparative Study

Ergosterol and ATP concentrations, microbial respiration and sporulation rates of aquatic hyphomycetes associated with leaves of Castanea sativa decomposing in a 5^th order stream were determined periodically over a period of 102 days in order to compare ergosterol and ATP as indicators of fungal biomass. ATP and ergosterol concentrations exhibited a significant positive correlation (F = 4.459, DF = 28, P < 0.001) during the first stages of leaf breakdown (until day 39), i.e., during periods of increasing fungal biomass. No correlation was found between ATP and ergosterol concentrations during later stages of decomposition (days 39 to 102). Respiration rates increased rapidly up to 0.525 mg O₂ h^–1 g^–1 AFDM during the first month and remained high until the end of the experiment. Sporulation rates peaked at day 9 (1069 conidia day^–1 mg^–1 AFDM) and decreased during later stages of decomposition. ATP‐to‐biomass conversion factors were determined for both fungi (0.59 μmol ATP g^–1 dry mass) and bacteria (1.30 μmol ATP g^–1 dry mass) collected from the stream and grown in the laboratory. Estimates of fungal biomass based on ATP concentrations were similar to those calculated from ergosterol concentrations during the first 39 days of breakdown. The results here presented suggest that ATP is a reliable method to quantify microbial biomass in streams and that the relative importance of bacteria increases at later stages of decomposition. (© 2009 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

Bimodal action of protons on ATP currents of rat PC12 cells

The mode of action of extracellular protons on ATP-gated P2X2 receptors remains controversial as either enhancement or depression of ATP-mediated currents has been reported. By investigating, at different pH, the electrophysiological effect of ATP on P2X2 receptors and complementing it with receptor modelling, the present study suggests a unified mechanism for both potentiation and inactivation of ATP receptors by protons. Our experiments on patch-clamped PC12 cells showed that, on the same cell, mild acidification potentiated currents induced by low ATP concentrations (<0.1 mM) and attenuated responses to high ATP concentrations (>1 mM) with emergence of current fading and rebound. To clarify the nature of the ATP/H+ interaction, we used the Ding and Sachs's "loop" receptor model which best describes the behavior of such receptors with two open states linked via one inactivated state. No effects by protons could be ascribed to H+-mediated open channel block. However, by assuming that protons facilitated binding of ATP to resting as well as open receptors, the model could closely replicate H+-induced potentiation of currents evoked by low ATP doses plus fading and rebound induced by high ATP doses. The latter phenomenon was due to receptor transition to the inactive state. The present data suggest that the high concentration of protons released with ATP (and catecholamines) from secretory vesicles may allow a dual action of H+ on P2X2 receptors. This condition might also occur on P2X2 receptors of central neurons exposed to low pH during ischemia.     

Gating of cystic fibrosis transmembrane conductance regulator chloride channels by adenosine triphosphate hydrolysis. Quantitative analysis of a cyclic gating scheme

Gating of the cystic fibrosis transmembrane conductance regulator (CFTR) involves a coordinated action of ATP on two nucleotide binding domains (NBD1 and NBD2). Previous studies using nonhydrolyzable ATP analogues and NBD mutant CFTR have suggested that nucleotide hydrolysis at NBD1 is required for opening of the channel, while hydrolysis of nucleotides at NBD2 controls channel closing. We studied ATP-dependent gating of CFTR in excised inside-out patches from stably transfected NIH3T3 cells. Single channel kinetics of CFTR gating at different [ATP] were analyzed. The closed time constant (tauc) decreased with increasing [ATP] to a minimum value of approximately 0.43 s at [ATP] >1.00 mM. The open time constant (tauo) increased with increasing [ATP] with a minimal tauo of approximately 260 ms. Kinetic analysis of K1250A-CFTR, a mutant that abolishes ATP hydrolysis at NBD2, reveals the presence of two open states. A short open state with a time constant of approximately 250 ms is dominant at low ATP concentrations (10 microM) and a much longer open state with a time constant of approximately 3 min is present at millimolar ATP. These data suggest that nucleotide binding and hydrolysis at NBD1 is coupled to channel opening and that the channel can close without nucleotide interaction with NBD2. A quantitative cyclic gating scheme with microscopic irreversibility was constructed based on the kinetic parameters derived from single-channel analysis. The estimated values of the kinetic parameters suggest that NBD1 and NBD2 are neither functionally nor biochemically equivalent.     

Optimization of adenosine 5′-triphosphate extraction for the measurement of␣acidogenic biomass utilizing whey wastewater

A set of experiments was carried out to maximize adenosine 5′-triphosphate (ATP) extraction efficiency from acidogenic culture using whey wastewater. ATP concentrations at different microbial concentrations increased linearly as microbial concentration decreased. More than 50% of ATP was extracted from the sample of 39 mg volatile suspended solids (VSS)/l compared to the sample of 2.8 g VSS/l. The ATP concentrations of the corresponding samples were 0.74±0.06 and 0.49±0.05 mg/l, respectively. For low VSS concentrations ranging from 39 to 92 mg/l, the extracted ATP concentration did not vary significantly at 0.73±0.01 mg ATP/l. Response surface methodology with a central composite in cube design for the experiments was used to locate the optimum for maximal ATP extraction with respect to boiling and bead beating treatments. The overall designed intervals were from 0 to 15 min and from 0 to 3 min for boiling and bead beating, respectively. The extracted ATP concentration ranged from 0.01 to 0.74 mg/l within the design boundary. The following is a partial cubic model where η is the concentration of ATP and x _k is the corresponding variable term (k=boiling time and bead beating time in order): η=0.629+0.035x ₁–0.818x ₂–0.002x ₁ x ₂–0.003x ₁ ² +0.254x ₂ ² +0.002x ₁ ² x ₂. This model successfully approximates the response of ATP concentration with respect to the boiling- and bead beating-time. The condition for maximal ATP extraction was 5.6 min boiling without bead beating. The maximal ATP concentration using the model was 0.74 mg/l, which was identical to the experimental value at optimum condition for ATP extraction.     

Microbial Biomass,Respiration, and Decomposition of Hura crepitans L. (Euphorbiaceae) Leaves in a Tropical Stream1

The processing of leaves in temperate streams has been the subject of numerous studies but equivalent tropical ecosystems have received little attention. We investigated leaf breakdown of a tropical tree species (Hura crepitans, Euphorbiaceae), in a tropical stream using leaf bags (0.5 mm mesh) over a period of 24 days. We followed the loss of mass and the changes in adenosine triphosphate (ATP) concentrations and respiration rates associated with the decomposing leaves. The breakdown rate was fast (k=?0.0672/d, k_d=?0.0031/degree‐day), with 81 percent loss of the initial mass within 24 days. This high rate was probably related to the stable and high water temperature (22°C) favoring strong biological activity. Respiration rates increased until day 16 (1.1 mg O₂/h/g AFDM), but maximum ATP concentrations were attained at day 9 (725 nmol ATP/g AFDM) when leaf mass remaining was 52 percent. To determine the relative importance of fungi and bacteria during leaf decomposition, ATP concentrations, and respiration rates were determined in samples treated with antibiotics, after incubation in the stream. The results of the samples treated with the antifungal or the bacterial antibiotic suggest a higher contribution of the fungal community for total microbial biomass and a higher contribution of the bacterial community for microbial respiration rates, especially during the later stages of leaf decomposition. However, these results should be analyzed with caution since both antibacterial and antifungal agents did not totally eliminate microbial activity and biomass.     

Occurrence and Ecological Significance of GTP in the Ocean and in Microbial Cells

A comparison between the ATP concentrations based on peak height light emission values (0 to 3 s) and integrated light flux determinations (15 to 75 s) for a variety of seawater samples revealed that the integrated method of light detection consistently yielded higher ATP concentrations, ranging from 1.38 to 2.35 times larger than the corresponding peak ATP values. A significant correlation (r = 0.923) was observed for a plot of ΔADP (i.e., integrated ATP - peak ATP) versus GTP + UTP, suggesting that the analytical interference on the ATP assay was the result of the presence of non-adenine nucleotide triphosphates. Size-fractionation studies revealed an enrichment of the non-adenine nucleotide triphosphates, relative to ATP, in the smallest size fraction analyzed (<10 μm). Investigations were conducted with 20 species of unicellular marine algae to determine their intracellular nucleotide concentrations, and these determinations were compared to the levels measured in lab cultures of the marine bacterium Serratia marinorubra. These results indicated that the intracellular GTP/ATP ratios in S. marinorubra increase in direct proportion to the rate of cell growth, and that the GTP/ATP ratios in bacteria are much greater than in growing algae, presumably due to the differences in rates of cellular biosynthesis. It is concluded that quantitative determinations of GTP/ATP ratios in environmental sample extracts may be useful for measuring microbial growth.     

Role of exercise-induced potassium fluxes underlying muscle fatigue: a brief review

The site of exercise-induced muscle fatigue is suggested to be the muscle membrane, which includes the sarcolemma and T-tubule membrane; the excitability of the membrane is dependent on the membrane potential. Significant potassium flux from the intracellular space of contracting muscle may decrease the membrane potential to half its resting value. This is true for isolated muscle preparations as well as for the whole body exercise in humans. Specific K+ channels have been identified, that may account for the intracellular K+ loss. Calcium-sensitive K+ channels open when intracellular Ca2+ concentrations increase, as during excitation. ATP-sensitive K+ channels may be involved but may open only at ATP concentrations well below those attained at exhaustion. However, ATP may be compartmentalized and only the membrane-bound ATP concentration may be of significance. Ca2+ accumulation and ATP depletion cause cell destruction; these changes induce an increased K+ conductance, which may inactivate the membrane and consequently prevent tension development. It is hypothesized that such a safety mechanism is identical to the fatigue mechanism.     

ATP as a biomass indicator in eight North Island Lakes, New Zealand

SUMMARY. 1. The relationship between microbial ATP and biomass- carbon in the near surface waters (0.5–5 m) of eight New Zealand lakes was studied to determine the constancy of the carbon:ATP ratio under natural growth conditions. Concentrations of microbial carbon were estimated indirectly from cell volume determinations.
2. The carbon:ATP ratio remained reasonably constant (interquartile range 248–291, n = 50), except during periods of nitrogen/phosphorus deficiency when carbon: ATP ratios increased to values greater than 400.
3. During periods of nitrogen/phosphorus sufficiency, corresponding estimates of microbial ATP and biomass-carbon were strongly correlated ( r =0.97, n =47) and related by the equation carbon = (287±20) ATP-(22±41) where carbon and ATP are expressed in mg m⁻³. From this relationship an average carbon:ATP ratio of 267 (SE=5) was calculated.
4. This ratio was not significantly affected by the relative proportions of bacterial and algal biomass in the surface water samples. However, because of the marked deviation of the carbon:ATP ratio during periods of nitrogen/phosphorus deficiency, the routine use of ATP as a biomass indicator is discouraged.     

Single channel properties of P2X2 purinoceptors

The single channel properties of cloned P2X2 purinoceptors expressed in human embryonic kidney (HEK) 293 cells and Xenopus oocytes were studied in outside-out patches. The mean single channel current-voltage relationship exhibited inward rectification in symmetric solutions with a chord conductance of approximately 30 pS at -100 mV in 145 mM NaCl. The channel open state exhibited fast flickering with significant power beyond 10 kHz. Conformational changes, not ionic blockade, appeared responsible for the flickering. The equilibrium constant of Na+ binding in the pore was approximately 150 mM at 0 mV and voltage dependent. The binding site appeared to be approximately 0.2 of the electrical distance from the extracellular surface. The mean channel current and the excess noise had the selectivity: K+ > Rb+ > Cs+ > Na+ > Li+. ATP increased the probability of being open (Po) to a maximum of 0.6 with an EC50 of 11.2 microM and a Hill coefficient of 2.3. Lowering extracellular pH enhanced the apparent affinity of the channel for ATP with a pKa of approximately 7.9, but did not cause a proton block of the open channel. High pH slowed the rise time to steps of ATP without affecting the fall time. The mean single channel amplitude was independent of pH, but the excess noise increased with decreasing pH. Kinetic analysis showed that ATP shortened the mean closed time but did not affect the mean open time. Maximum likelihood kinetic fitting of idealized single channel currents at different ATP concentrations produced a model with four sequential closed states (three binding steps) branching to two open states that converged on a final closed state. The ATP association rates increased with the sequential binding of ATP showing that the binding sites are not independent, but positively cooperative. Partially liganded channels do not appear to open. The predicted Po vs. ATP concentration closely matches the single channel current dose-response curve.     

ATP regulation of type 1 inositol 1,4,5-trisphosphate receptor channel gating by allosteric tuning of Ca(2+) activation.

Inositol 1,4,5-trisphosphate (InsP(3)) mobilizes intracellular Ca(2+) by binding to its receptor (InsP(3)R), an endoplasmic reticulum-localized Ca(2+) release channel. Patch clamp electrophysiology of Xenopus oocyte nuclei was used to study the effects of cytoplasmic ATP concentration on the cytoplasmic Ca(2+) ([Ca(2+)](i)) dependence of single type 1 InsP(3)R channels in native endoplasmic reticulum membrane. Cytoplasmic ATP free-acid ([ATP](i)), but not the MgATP complex, activated gating of the InsP(3)-liganded InsP(3)R, by stabilizing open channel state(s) and destabilizing the closed state(s). Activation was associated with a reduction of the half-maximal activating [Ca(2+)](i) from 500 +/- 50 nM in 0 [ATP](i) to 29 +/- 4 nM in 9.5 mM [ATP](i), with apparent ATP affinity = 0.27 +/- 0.04 mM, similar to in vivo concentrations. In contrast, ATP was without effect on maximum open probability or the Hill coefficient for Ca(2+) activation. Thus, ATP enhances gating of the InsP(3)R by allosteric regulation of the Ca(2+) sensitivity of the Ca(2+) activation sites of the channel. By regulating the Ca(2+)-induced Ca(2+) release properties of the InsP(3)R, ATP may play an important role in shaping cytoplasmic Ca(2+) signals, possibly linking cell metabolic state to important Ca(2+)-dependent processes.     

ATP levels in algal cells as influenced by environmental conditions

The cellular content of adenosine triphosphate (ATP) relativeto cell size and cellular organic carbon has been investigatedin 30 different algal cultures representing 7 phyla. Duringexponential growth in batch culture, cellular contents of ATPremained at fairly uniform levels in all these unicellular algaeand averaged 0.35% of the cellular organic carbon content. Duringextreme nitrogen or phosphorus deficiency the cellular levelsof ATP decreased to 20–50% of that found in exponentially-growingcells, but these percentages may be low due to detrital carbonin the senescent cultures. The steady state levels of ATP in cells were similar in lightor in dark, although ATP concentrations fluctuated for a fewminutes upon any sudden change in light conditions. When thelight was turned on there was a rapid increase in ATP levels,followed by a slow decrease; when the light was turned off,there was a rapid fall in cellular ATP levels, which then rosewithin a few minutes to achieve the steady state concentration.The cellular concentrations of ATP in these algae and in othermicrobial groups are discussed relative to studies where ATPdeterminations are used to estimate microbial biomass. (Received June 2, 1970; )     

The kinetic and physical basis of K(ATP) channel gating: toward a unified molecular understanding

K(ATP) channels can be formed from Kir6.2 subunits with or without SUR1. The open-state stability of K(ATP) channels can be increased or reduced by mutations throughout the Kir6.2 subunit, and is increased by application of PIP(2) to the cytoplasmic membrane. Increase of open-state stability is manifested as an increase in the channel open probability in the absence of ATP (Po(zero)) and a correlated decrease in sensitivity to inhibition by ATP. Single channel lifetime analyses were performed on wild-type and I154C mutant channels expressed with, and without, SUR1. Channel kinetics include a single, invariant, open duration; an invariant, brief, closed duration; and longer closed events consisting of a "mixture of exponentials," which are prolonged in ATP and shortened after PIP(2) treatment. The steady-state and kinetic data cannot be accounted for by assuming that ATP binds to the channel and causes a gate to close. Rather, we show that they can be explained by models that assume the following regarding the gating behavior: 1) the channel undergoes ATP-insensitive transitions from the open state to a short closed state (C(f)) and to a longer-lived closed state (C(0)); 2) the C(0) state is destabilized in the presence of SUR1; and 3) ATP can access this C(0) state, stabilizing it and thereby inhibiting macroscopic currents. The effect of PIP(2) and mutations that stabilize the open state is then to shift the equilibrium of the "critical transition" from the open state to the ATP-accessible C(0) state toward the O state, reducing accessibility of the C(0) state, and hence reducing ATP sensitivity.