Expression of the human Menkes ATPase in Xenopus laevis oocytes.
Menkes disease is an X-linked disorder of copper metabolism that is usually fatal. The affected gene has recently been cloned and encodes one of the two human copper ATPases. If the Menkes ATPase is defective, copper is trapped in the intestinal mucosa, leading to systemic copper deficiency. In order to study copper transport by this ATPase and the effects of disease mutations on its function, we developed a Xenopus laevis oocyte expression system. Wild-type Menkes ATPase cDNA and a fusion of this gene with the green fluorescent protein (GFP) gene was transcribed in vitro and the mRNA injected into oocytes. Expression in oocytes was analyzed by Western blotting and fluorescence microscopy. The Menkes ATPase-GFP chimera appeared to localize primarily to the plasma membrane as assessed by confocal microscopy. This system should thus provide an interesting new tool to study the function of the Menkes ATPase.
Bissig
K D
KD
Department of Clinical Pharmacology, University of Berne, Switzerland.
La Fontaine
S
S
Mercer
J F
JF
Solioz
M
M
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000818:Animals; D019677:Artificial Gene Fusion; D002352:Carrier Proteins / Q000378:metabolism*; D027682:Cation Transport Proteins; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D003300:Copper / Q000172:deficiency*; D000073840:Copper-Transporting ATPases; D005260:Female; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D009865:Oocytes / Q000201:enzymology* / Q000648:ultrastructure; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014982:Xenopus laevis
D002352:Carrier Proteins; D027682:Cation Transport Proteins; D008164:Luminescent Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D003300:Copper; D000251:Adenosine Triphosphatases; C081184:ATP7A protein, human; D000073840:Copper-Transporting ATPases