Bong-Heuy Cho

The role of potassium in charge compensation for sucrose-proton-symport by cotyledons of Ricinus communis

Abstract The addition of sucrose to cotyledons of Ricinus communis caused a transient uptake of protons and, simultaneously, a transient efflux of potassium ions. The stoichiometry was up to approx. 0.3 protons taken up per sucrose and up to 0.6 potassium ions released per sucrose. The presence of lipophilic cations or anions strongly reduced the potassium efflux with less change of the proton/sucrose stoichiometry. It is concluded that the uptake of protons with sucrose is mechanistically coupled by the sucrose transport system whereas the potassium efflux occurs passively by some other path and is caused by membrane potential depolarisation. The sucrose uptake system is therefore an electrogenic proton symport system, and the efflux of potassium only serves for charge compensation.

The amino acid transport systems of the autotrophically grown green alga chlorella

Abstract The kinetic analysis of l -amino acid uptake by the green alga Chlorella revealed at least seven different uptake systems to be present in cells grown autotrophically with nitrate as nitrogen source. There is a ‘general system’ which transports most neutral and acidic amino acids, a system for short-chain neutral amino acids including proline, a system for basic amino acids including histidine, a special system for acidic amino acids, and specific systems for methionine, glutamine and threonine. The ‘general system’ is possibly the same as that which can be stimulated by incubation of cells in glucose plus ammonium (Sauer, N. (1984) Planta 161, 425–431). The incubation of Chlorella in glucose induces the increased synthesis of six amino acid uptake systems, namely the above-mentioned system for short-chain neutral amino acids, a threonine system, a methionine system, and a glutamine system. These results indicate that the uptake of l -amino acids by the green alga Chlorella is as complex as in other free-living organisms such as bacteria or yeast. The small number of amino acid uptake systems found in cells of higher plants, i.e. two or three, seems therefore to be a consequence of integration of the cells in a tissue supplying a relatively constant environment, and not a consequence of autotrophic growth on mineral carbon and mineral nitrogen.

Mechanism of proline uptake by Chlorella vulgaris

An amino acid uptake system specific for glycine, alanine, serine and proline was induced by glucose in Chlorella vulgaris. The uptake system translocated the zwitterionic form of the amino acid. There was more than 100-fold accumulation which indicated a coupling to metabolic energy. The depolarization of the membrane potential during proline uptake and the sensitivity of its uptake rate to the membrane potential point to coupling with an ion flow. Inhibitors of plasmalemma-bound H+-ATPase inhibit proline uptake. These data are interpreted to mean that proline is taken up as a proton symport. In some Chlorella strains the proline-coupled H+ uptake could be measured with electrodes, but not in Chlorella vulgaris. There is evidence that the transport of amino acids rapidly stimulates the proton-translocating ATPase of Chlorella vulgaris, so that the proline-coupled proton uptake is immediately neutralized.

Regulation of nitrate uptake by glucose in chlorella

Abstract The rate of nitrate uptake by Chlorella vulgaris was stimulated 5-fold when the cells were preincubated with glucose. The effect of glucose was probably by induction since the timecourse of stimulation revealed a lag period of approx. 30 min. Preincubation with 6-deoxyglucose caused no stimulation. The presence of glucose (or ammonium) during nitrate uptake had no influence on the uptake rate. The transport of nitrate was accompanied by H+ symport in a stoichiometry of 1:1.

Charge and acidity compensation during proton-sugar symport in Chlorella: The H(+)-ATPase does not fully compensate for the sugar-coupled proton influx.

This study was undertaken in order to demonstrate the extent to which the activity of the plasmalemma H+-ATPase compensates for the charge and acidity flow caused by the sugar-proton symport in cells of chlorella vulgaris Beij.. Detailed analysis of H+ and K+ fluxes from and into the medium together with measurements of respiration, cytoplasmic pH, and cellular ATP-levels indicate three consecutive phases after the onset of H+ symport. Phase 1 occurred immediately after addition of sugar, with an uptake of H+ by the hexoseproton symport and charge compensation by K+ loss from the cells and, to a smaller degree, by loss of another ion, probably a divalent cation. This phase coincided with strong membrane depolarization. Phase 2 started approximately 5 s after addition of sugar, when the acceleration of the H+-ATPase caused a slow-down of the K+ efflux, a decrease in the cellular ATP level and an increase in respiration. The increased respiration was most probably responsible for a pronounced net acidification of the medium. This phase was inhibited in deuterium oxide. In phase 3, finally, a slow rate of net H+ uptake and K+ loss was established for several further minutes, together with a slight depolarization of the membrane. There was hardly any pH change in the cytoplasm, because the cytoplasmic buffering capacity was high enough to stabilize the pH for several minutes despite the net H+ fluxes. The quantitative participation of the several phases of H+ and K+ flow depended on the pH of the medium, the ambient Ca2+ concentration, and the metabolic fate of the transported sugar. The results indicate that the activity of the H+-ATPase never fully compensated for H+ uptake by the sugar-symport system, because at least 10% of symport-caused charge inflow was compensated for by K+ efflux. The restoration of pH in the cytoplasm and in the medium was probably achieved by metabolic reactions connected to increased glycolysis and respiration.

Mechanism of arginine transport in Chlorella

The incubation of Chlorella cells with glucose causes the induction of an uptake system, which is specific for the basic amino acids arginine and lysine. Both amino acids are taken up in the positively charged form and with high affinity (Kmvalues 2 μM and 7 μM, respectively). The transport of arginine depolarizes the membrane by 20–30 mV. The charge compensation is achieved within a few seconds after arginine addition by the proton pump, later on K+ efflux serves for charge compensation. No evidence for arginine-proton symport was found, neither by inhibitor studies nor by use of other Chlorella strains which have a slower-responding proton pump. The accumulation of arginine is appreciably higher than it should be according to the thermodynamic force of the membrane potential. There is, however, some evidence that a large proportion of arginine is trapped by intracellular compartments and is therefore not in equilibrium with the outside arginine.

The hexose uptake system of Chlorella : Is it a proton symport or a hydroxyl ion antiport system?

2‐Amino‐2‐deoxyglucose is taken up by the hexose transport system of Chlorella. The uptake of the neutral sugar proceeds together with stoichiometric alkalization of the medium. The uptake of the positively charged sugar (i.e., with a protonated amino group) proceeds as uniport and is not accompanied by net proton uptake but by charge compensating exit of proton and potassium. Thus the proton at the amino group replaces the cotransported ion, which therefore has to be a proton and not a hydroxyl ion. A depolarization of the membrane from − 130 mV to − 70 mV increased the K m‐value 3‐fold without change of V max. It is concluded that neutral sugar and proton are bound in close vicinity at a site in the middle of the membrane and that transport of sugar and proton occurs simultaneously.

Comparison of suspension cells and cotyledons of ricinus with respect to sugar uptake.

Suspension cells and cotyledons of Ricinus communis were compared as to their uptake properties for sugar and amino acids to reveal whether the previously reported sucrose-specificity of the cotyledon is a specific feature of the cotyledon or of the Ricinus cell in general. The experiments show that suspension cells have a higher hexose uptake activity at low sugar concentration than cotyledons, whereas sucrose cannot be taken up by suspension cells unless it is first hydrolyzed. Amino acids are taken up by suspension cells and by the cotyledons. It is concluded that the highly specific uptake of sucrose without hydrolysis is a special feature of certain specialized cells of the cotyledon, probably the phloem.