PMID:21621506

Citation	Leitenmaier, B, Witt, A, Witzke, A, Stemke, A, Meyer-Klaucke, W, Kroneck, PM and Küpper, H (2011) Biochemical and biophysical characterisation yields insights into the mechanism of a Cd/Zn transporting ATPase purified from the hyperaccumulator plant Thlaspi caerulescens. Biochim. Biophys. Acta 1808:2591-9
Abstract	TcHMA4 (GenBank no. AJ567384), a Cd/Zn transporting ATPase of the P(1B)-type (=CPx-type) was isolated and purified from roots of the Cd/Zn hyperaccumulator Thlaspi caerulescens. Optimisation of the purification protocol, based on binding of the natural C-terminal His-tag of the protein to a Ni-IDA metal affinity column, yielded pure, active TcHMA4 in quantities sufficient for its biochemical and biophysical characterisation with various techniques. TcHMA4 showed activity with Cu(2+), Zn(2+) and Cd(2+) under various concentrations (tested from 30nM to 10μM), and all three metal ions activated the ATPase at a concentration of 0.3μM. Notably, the enzyme worked best at rather high temperatures, with an activity optimum at 42°C. Arrhenius plots yielded interesting differences in activation energy. In the presence of zinc it remained constant (E(A)=38kJ⋅mol(-1)) over the whole concentration range while it increased from 17 to 42kJ⋅mol(-1) with rising copper concentration and decreased from 39 to 23kJ⋅mol(-1) with rising cadmium concentration. According to EXAFS the TcHMA4 appeared to bind Cd(2+) mainly by thiolate sulphur from cysteine, and not by imidazole nitrogen from histidine.
Links	PubMed Online version:10.1016/j.bbamem.2011.05.010
Keywords	Adenosine Triphosphatases/isolation & purification; Adenosine Triphosphatases/metabolism; Biophysics; Cadmium/metabolism; Electrophoresis, Polyacrylamide Gel; Spectrophotometry, Atomic; Zinc/metabolism
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
THLCA:Q70LF4		GO:0046873: metal ion transmembrane transporter activity	ECO:0000314:		F		Fig 2. shows that the ATPase activity of the transporter is activated by Cadmium, copper, and zinc	complete CACAO 4767

References

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