Functional reconstitution of Arabidopsis thaliana plant uncoupling mitochondrial protein (AtPUMP1) expressed in Escherichia coli.
The Arabidopsis thaliana uncoupling protein (UCP) gene was expressed in Escherichia coli and isolated protein reconstituted into liposomes. Linoleic acid-induced H+ fluxes were sensitive to purine nucleotide inhibition with an apparent K(i) (in mM) of 0.8 (GDP), 0.85 (ATP), 0.98 (GTP), and 1.41 (ADP); the inhibition was pH-dependent. Kinetics of AtPUMP1-mediated H+ fluxes were determined for lauric, myristic, palmitic, oleic, linoleic, and linolenic acids. Properties of recombinant AtPUMP1 indicate that it represents a plant counterpart of animal UCP2 or UCP3. This work brings the functional and genetic approaches together for the first time, providing strong support that AtPUMP1 is truly an UCP.
Borecký
J
J
Department of Membrane Transport Biophysics, Institute of Physiology, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
Maia
I G
IG
Costa
A D
AD
Jezek
P
P
Chaimovich
H
H
de Andrade
P B
PB
Vercesi
A E
AE
Arruda
P
P
D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D017360:Arabidopsis / Q000737:chemistry*; D029681:Arabidopsis Proteins; D015153:Blotting, Western; D002352:Carrier Proteins / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism*; D005822:Genetic Vectors; D006153:Guanosine Diphosphate / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D006859:Hydrogen / Q000378:metabolism; D007473:Ion Channels; D007700:Kinetics; D019787:Linoleic Acid / Q000494:pharmacology; D008565:Membrane Proteins / Q000378:metabolism*; D026901:Membrane Transport Proteins; D024101:Mitochondrial Proteins; D000071245:Mitochondrial Uncoupling Proteins; D009841:Oligonucleotides / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011506:Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D000071256:Uncoupling Protein 1; D000071246:Uncoupling Protein 2; D000071247:Uncoupling Protein 3
D029681:Arabidopsis Proteins; D002352:Carrier Proteins; D007473:Ion Channels; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D024101:Mitochondrial Proteins; D000071245:Mitochondrial Uncoupling Proteins; D009841:Oligonucleotides; C000604370:PUMP2 protein, Arabidopsis; D011506:Proteins; D011994:Recombinant Proteins; D000071256:Uncoupling Protein 1; D000071246:Uncoupling Protein 2; D000071247:Uncoupling Protein 3; D006153:Guanosine Diphosphate; D000244:Adenosine Diphosphate; D006859:Hydrogen; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D019787:Linoleic Acid