Functional reconstitution of an ATP-driven Ca sup 2+ -transport system from the plasma membrane of Commelina communis L

Journal Article · Mon Oct 01 00:00:00 EDT 1990 · Plant Physiology; (USA)

DOI:https://doi.org/10.1104/pp.94.2.634· OSTI ID:5548892

Graef, P; Weiler, E W ^[1]

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1. Ruhr-Universitaet Bochum (West Germany)

The protein(s) that constitute(s) the ATP-driven Ca{sup 2+}-translocator of plasma membrane enriched vesicles obtained by aqueous two-phase partitioning from leaves of Commelina communis L. has/have been solubilized and reincorporated into tightly sealed liposomes. The reconstituted Ca{sup 2+}-transport system was studied using ATP-driven {sup 45}Ca{sup 2+} import into the proteoliposomes as a measure of activity. The detergent, 3- ((3-cholamidopropyl) dimethylammonio) -1-propane-sulfonate proved to be the most suitable and was used at 10 millimolar concentration, i.e. just above its critical micellar concentration. The presence of additional phospholipid and ATP improved the solubilization and/or reconstitution. The characteristics of the reconstituted system were similar to those of the plasma membrane-bound activity, including the apparent K{sub m} for Ca{sup 2+} inhibition by relatively high levels of vanadate and lacking response to added calmodulin. The reconstituted transport system was very strongly inhibited by erythrosine B and had a low apparent K{sub m} for ATP levels of the Ca{sup 2+}-ionophore A 23187 instantaneously discharged 90% of the Ca{sup 2+} associated with the vesicles, proving that it had been accumulated in the intravesicular volume in soluble, freely exchangeable form. Ca{sup 2+}-transport in the reconstituted system was thus primary active, through a Ca{sup 2+}-translocating ATPase.

• OSTI ID: 5548892
• Journal Information: Plant Physiology; (USA), Vol. 94:2; ISSN 0032-0889
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
ATP-ASE
BIOCHEMICAL REACTION KINETICS
CALCIUM COMPOUNDS
MEMBRANE TRANSPORT
CALCIUM 45
CELL MEMBRANES
PLANTS
TRACER TECHNIQUES
ACID ANHYDRASES
ALKALINE EARTH ISOTOPES
ALKALINE EARTH METAL COMPOUNDS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
CALCIUM ISOTOPES
CELL CONSTITUENTS
DAYS LIVING RADIOISOTOPES
ENZYMES
EVEN-ODD NUCLEI
HYDROLASES
INTERMEDIATE MASS NUCLEI
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
MEMBRANES
NUCLEI
PHOSPHOHYDROLASES
RADIOISOTOPES
REACTION KINETICS
551001* - Physiological Systems- Tracer Techniques