The kinetics of facilitated diffusion followed by enzymatic conversion of the substrate

Kuile, Benno H. ter; Cook, Michael

doi:10.1016/0005-2736(94)90158-9

Cited by 18 publications

(16 citation statements)

References 14 publications

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“…An important implication of the results presented in this paper is that a 5-s transport experiment does not, under some circumstances, adequately report the true kinetics of the glucose transporter in S. cerevisiae. Indeed, based on modelling studies, it has been demonstrated that apparent changes in kinetics of a transport process can be induced by changes in the subsequent metabolism (ter Kuile et al, 1994;Fuhrmann and Volker, 1992). Thus, relatively long-term (5 s) transport experiments may lead to erroneous conclusions concerning the regulation of this process, especially in mutants affected in metabolism.…”

Section: Discussionmentioning

confidence: 99%

High‐affinity Glucose Uptake in Saccharomyces cerevisiae is not Dependent on the Presence of Glucose‐Phosphorylating Enzymes

SMITS¹,

SMITS²,

POSTMA³

et al. 1996

Yeast

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Glucose uptake in Saccharomyces cerevisiae is believed to consist of two kinetically distinguishable components, the affinity of which is modulated during growth on glucose. It has been reported that triple hexose-kinase deletion mutants do not exhibit high-affinity glucose uptake. This raises the question of whether and how high-affinity glucose uptake is related to the presence of glucose-phosphorylating enzymes. In this study the kinetics of glucose uptake in both wild-type cells and cells of hexose-kinase deletion mutants, grown on either glycerol or galactose, were determined using a rapid-uptake method. In wild-type cells glucose uptake measured over either 5 s or 200 ms exhibited high affinity. In contrast, in cells of hexose-kinase deletion mutants the apparent affinity of glucose uptake was dependent on the time scale during which uptake was measured. Measurements on the 5-s scale showed apparent low-affinity uptake whereas measurements on the 200-ms scale showed high-affinity uptake. The affinity and maximal rate of the latter were comparable to those in wild-type cells. Using a simple model for a symmetrical facilitator, it was possible to simulate the experimentally determined relation between apparent affinity and the time scale used. The results suggest that high-affinity glucose transport is not necessarily dependent on the presence of glucose-phosphorylating enzymes. Apparent low-affinity uptake kinetics can arise as a consequence of an insufficient rate of removal of intracellular free glucose by phosphorylation. This study underlines the need to differentiate between influences of the translocator and of metabolism on the apparent kinetics of sugar uptake in yeast.

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Section: Discussionmentioning

confidence: 99%

High‐affinity Glucose Uptake in Saccharomyces cerevisiae is not Dependent on the Presence of Glucose‐Phosphorylating Enzymes

SMITS¹,

SMITS²,

POSTMA³

et al. 1996

Yeast

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“…However, the possible involvement of a simultaneous facilitated diffusion process could not be completely ruled out as the low molecular mass of the ferric iron dicitrate complex of 443 Da may allow its diffusion through the PBM, as observed in E. coli for the cloned iron uptake system of Serratia marcescens [29]. The simultaneous existence of these two processes could explain the variability in the Km determination [30].…”

Section: Discussionmentioning

confidence: 99%

Uptake of iron by symbiosomes and bacteroids from soybean nodules

1995

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Symbiosomes and bacteroids isolated from soybean nodules are able to take up the iron--citrate complex. The kinetics are characterized by initial high rates of iron internalization, and ATPase inhibitors significantly lower the uptake. This is consistent with an energy-dependent process on both membranes, although the involvement of a simultaneous facilitated diffusion can not be completely ruled out. Citrate alone is poorly absorbed by symbiosomes; this uptake is greatly enhanced by addition of iron. Iron-citrate was found both in the nodule cytosol and in the bacteroids. These results provide the first experimental evidence for the existence, at least in young nodules, of an important iron trafficking system from the plant host cell to the microsymbiont, through the peribacteroid membrane.Key words': Iron uptake; Peribacteroid membrane; Citrate; Nitrogen fixation; Root nodule are always surrounded by a membrane of plasmic origin [1]; the so-called peribacteroid membrane (PBM) isolates the microbial symbiont from the cytoplasm of the host cell to form the symbiosome, delimiting a peribacteroid space (PBS). Thus, the symbiosome constitutes the entire functional entity suitable for exchange studies between the two partners. There are several lines of evidence that PBM exhibits a restricted permeability. Till now, only dicarboxylate transport activities [10,11] and glucose permeability [11] have been observed.In this work, we characterize for the first time the uptake of iron by purified symbiosomes from soybean nodules. The role of citrate in this uptake is highlighted and the involvement of these processes in functioning nodules is discussed. Materials and methods ~. IntroductionPlants of the family Leguminosae are, together with rhizobial microsymbionts, able to form root nodules capable of reducing atmospheric dinitrogen to ammonia [1]. Iron metabolism is of ~articular importance in nodules since this metal is a constitu-,'nt of key proteins such as nitrogenase and leghemoglobin Lb). Because of this, heine and heme proteins are of special nterest because the cytochrome content changes both qualitaively and quantitatively during bacterial differentiation into ~ymbiotic bacteroids [2]. In addition, there is an important need "or heme in the synthesis of leghemoglobin in vivo, which hakes up to 20% of the plant protein mass in the nodule cytosol 2]. Based on the failure to detect heine biosynthetic enzyme lctivities in the plant cytosol of root nodules, it was first projgosed that the heme for Lb is exclusively synthesized by the ~'hizobial microsymbiont [3,4]. More recent investigations have ,hown that the levels of several plant enzymes in the heine biosynthesis pathway are increased in nodules, compared to roots, indicating that the plant may also contribute to heme formation [5][6][7][8]. Although the symbiont that carries out Lb taeme synthesis remains uncertain [9], the fact remains that, to ~ynthesize their cytochromes and other redox components organized as a branched system [2], bacteroids need ...

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“…brzlcei (Ter Kuile & Opperdoes, 1992~). The biphasic uptake kinetics of the type observed for glucose uptake by T. vaginalis clearly indicate facilitated diffusion as transport mechanism followed by enzymic conversion of the substrate (Ter Kuile & Cook, 1994) and thus induction of an active carrier can be ruled out. The facilitated diffusion carrier cannot concentrate glucose against the gradient (Stein, 1986), but an increase in the number of carriers will lead to increased uptake if the capacity of the subsequent enzymic pathway is also increased.…”

Section: Adaptation To Substrate Availabilitymentioning

confidence: 88%

Adaptation of the carbon metabolism of Trichomonas vaginalis to the nature and availability of the carbon source

Kuile

1994

Microbiology

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The anaerobic parasitic protist Trichomonas vaginalis was adapted in chemostats to eight different conditions defined by different growth rates and carbon regimes. Glucose or maltose was used as carbon and energy source. Cells cultured under well-defined steady states were tested in short-term experiments. The kinetics of glucose and maltose uptake were determined and their glucokinase and a-glucosidase activities were measured. Uptake in 20 min was measured with radiolabelled glucose and maltose, rather than analogues, using the silicone oil centrifugation technique. Hence, the accumulated label represents both transport and metabolic activity. The total uptake of glucose was highest in organisms that had been starved for glucose during growth. The kinetics of glucose uptake can be understood b y assuming rate-limitation by transport across the plasma membrane at low external concentrations and by the subsequent metabolism a t concentrations exceeding a cross-over value. The specific glucokinase activity correlated in only four out of eight cases with the saturation uptake. The kinetics of maltose uptake indicated rate-limitation a t low maltose concentrations by a diffusion-limited step and a t higher levels by metabolic steps. The uptake of maltose was primarily affected by the growth rate during culture, the highest growth rates resulting in most uptake. Maltose uptake was determined only partially by the cellular a-glucosidase activity. The activities of both transport and metabolic enzymes changed due to the culture conditions suggesting that the control over glucose and maltose metabolism is shared by several steps in the pathway.

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The kinetics of facilitated diffusion followed by enzymatic conversion of the substrate

Cited by 18 publications

References 14 publications

High‐affinity Glucose Uptake in Saccharomyces cerevisiae is not Dependent on the Presence of Glucose‐Phosphorylating Enzymes

High‐affinity Glucose Uptake in Saccharomyces cerevisiae is not Dependent on the Presence of Glucose‐Phosphorylating Enzymes

Uptake of iron by symbiosomes and bacteroids from soybean nodules

Adaptation of the carbon metabolism of Trichomonas vaginalis to the nature and availability of the carbon source

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