Functional reconstitution of Arabidopsis thaliana plant uncoupling mitochondrial protein (At PUMP1) expressed in Escherichia coli (original) (raw)
Related papers
Journal of Bioenergetics and Biomembranes, 2000
The presence of plant-uncoupling mitochondrial protein (PUMP), previously described by , was screened in mitochondria of various organs or tissues of several plant species. This was done functionally, by monitoring purine nucleotide-sensitive linoleic acid-induced uncoupling, or by Western blots. The following findings were established: (1) PUMP was found in most of the higher plants tested; (2) since ATP inhibition of linoleic acidinduced membrane potential decrease varied, PUMP content might differ in different plant tissues, as observed with mitochondria from maize roots, maize seeds, spinach leaves, wheat shoots, carrot roots, cauliflower, broccoli, maize shoots, turnip root, and potato calli. Western blots also indicated PUMP presence in oat shoots, carnation petals, onion bulbs, red beet root, green cabbage, and Sedum leaves. (3) PUMP was not detected in mushrooms. We conclude that PUMP is likely present in the mitochondria of organs and tissues of all higher plants.
AtPUMP: an Arabidopsis gene encoding a plant uncoupling mitochondrial protein
FEBS Letters, 1998
A cDNA clone (AtPUMP) encoding a plant uncoupling mitochondrial protein was isolated from Arabidopsis thaliana. The cDNA contains an open reading frame of 921 nucleotides encoding 306 amino acids (predicted molecular weight 32 708). The predicted polypeptide is 81% identical and 89% similar to the potato UCP-like protein, and includes an energy transfer protein motif common to mitochondrial transporters. The AtPUMP gene exists as a single copy in the Arabidopsis genome. The corresponding transcript was expressed in all tissues and was strongly induced by cold treatment. We suggest that the putative AtPUMP protein may play a role in heat-requiring physiological events in Arabidopsis.
Mutational analysis of Arabidopsis thaliana plant uncoupling mitochondrial protein
Biochimica et Biophysica Acta (BBA) - Bioenergetics, 2007
In this study, point mutations were introduced in plant uncoupling mitochondrial protein AtUCP1, a typical member of the plant uncoupling protein (UCP) gene subfamily, in amino acid residues Lys147, Arg155 and Tyr269, located inside the so-called UCP-signatures, and in two more residues, Cys28 and His83, specific for plant UCPs. The effects of amino acid replacements on AtUCP1 biochemical properties were examined using reconstituted proteoliposomes. Residue Arg155 appears to be crucial for AtUCP1 affinity to linoleic acid (LA) whereas His83 plays an important role in AtUCP1 transport activity. Residues Cys28, Lys147, and also Tyr269 are probably essential for correct protein function, as their substitutions affected either the AtUCP1 affinity to LA and its transport activity, or sensitivity to inhibitors (purine nucleotides). Interestingly, Cys28 substitution reduced ATP inhibitory effect on AtUCP1, while Tyr269Phe mutant exhibited 2.8-fold increase in sensitivity to ATP, in accordance with the reverse mutation Phe267Tyr of mammalian UCP1.
AtUCP2: a Novel Isoform of the Mitochondrial Uncoupling Protein of Arabidopsis thaliana
Plant and Cell Physiology, 1999
Mitochondrial uncoupling proteins (UCPs) play a central role in adaptive thermogenesis in mammals. The UCPs dissipate the proton gradient formed through respiration without ATP synthesis, and the freed energy is readily converted to heat, helping the animals to maintain their body temperature in cold environments. Recently, it was found that UCPs also function in plant mitochondria. Subsequently, a cDNA clone encoding a UCP in potato was isolated. Whereas the UCP gene constitutes a multigene family in mammals, only a single cDNA sequence has been reported so far for the potato UCP. Moreover, it has been recently suggested that Arabidopsis has only a single nuclear gene for UCP. Here we report the existence of another UCP gene in the Arabidopsis genome, showing for the first time the occurrence of a multigene family for the protein in higher plants. A cDNA analysis of this gene showed that the novel isoform possesses all typical features reported for known UCPs. However, the new gene, unlike the other gene in Arabidopsis and the gene in potato, did not appear to respond to low temperature.
PLANT UNCOUPLING MITOCHONDRIAL PROTEINS
Annual Review of Plant Biology, 2006
Uncoupling proteins (UCPs) are membrane proteins that mediate purine nucleotide-sensitive free fatty acid-activated H + flux through the inner mitochondrial membrane. After the discovery of UCP in higher plants in 1995, it was acknowledged that these proteins are widely distributed in eukaryotic organisms. The widespread presence of UCPs in eukaryotes implies that these proteins may have functions other than thermogenesis. In this review, we describe the current knowledge of plant UCPs, including their discovery, biochemical properties, distribution, gene family, gene expression profiles, regulation of gene expression, and evolutionary aspects. Expression analyses and functional studies on the plant UCPs under normal and stressful conditions suggest that UCPs regulate energy metabolism in the cellular responses to stress through regulation of the electrochemical proton potential ( μ H +) and production of reactive oxygen species.
Proteomic Approach to Identify Novel Mitochondrial Proteins in Arabidopsis
Plant Physiology, 2001
An Arabidopsis mitochondrial proteome project was started for a comprehensive investigation of mitochondrial functions in plants. Mitochondria were prepared from Arabidopsis stems and leaves or from Arabidopsis suspension cell cultures, and the purity of the generated fractions was tested by the resolution of organellar protein complexes applying two-dimensional blue-native/N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine (Tricine) sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The Arabidopsis mitochondrial proteome was analyzed by two-dimensional isoelectric focusing/ Tricine sodium dodecyl sulfate-polyacrylamide gel electrophoresis and 650 different proteins in a pI range of pH 3 to 10 were separated on single gels. Solubilization conditions, pH gradients for isoelectric focusing, and gel staining procedures were varied, and the number of separable proteins increased to about 800. Fifty-two protein spots were identified by immunoblotting, direct protein sequencing, and...