Chaperone activity of recombinant maize chloroplast protein synthesis elongation factor, EF-Tu.
The protein synthesis elongation factor, EF-Tu, is a protein that carries aminoacyl-tRNA to the A-site of the ribosome during the elongation phase of protein synthesis. In maize (Zea mays L) this protein has been implicated in heat tolerance, and it has been hypothesized that EF-Tu confers heat tolerance by acting as a molecular chaperone and protecting heat-labile proteins from thermal aggregation and inactivation. In this study we investigated the effect of the recombinant precursor of maize EF-Tu (pre-EF-Tu) on thermal aggregation and inactivation of the heat-labile proteins, citrate synthase and malate dehydrogenase. The recombinant pre-EF-Tu was purified from Escherichia coli expressing this protein, and mass spectrometry confirmed that the isolated protein was indeed maize EF-Tu. The purified protein was capable of binding GDP (indicative of protein activity) and was stable at 45 degrees C, the highest temperature used in this study to test this protein for possible chaperone activity. Importantly, the recombinant maize pre-EF-Tu displayed chaperone activity. It protected citrate synthase and malate dehydrogenase from thermal aggregation and inactivation. To our knowledge, this is the first observation of chaperone activity by a plant/eukaryotic pre-EF-Tu protein. The results of this study support the hypothesis that maize EF-Tu plays a role in heat tolerance by acting as a molecular chaperone and protecting chloroplast proteins from thermal aggregation and inactivation.
Rao
Damodara
D
Department of Biology, University of South Dakota, School of Medicine, Vermillion, SD 57069, USA.
Momcilovic
Ivana
I
Kobayashi
Satoru
S
Callegari
Eduardo
E
Ristic
Zoran
Z
D000595:Amino Acid Sequence; D015153:Blotting, Western; D002736:Chloroplasts / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000378:metabolism; D007526:Isoelectric Point; D018832:Molecular Chaperones / Q000378:metabolism*; D008970:Molecular Weight; D010444:Peptide Elongation Factor Tu / Q000378:metabolism*; D011485:Protein Binding; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism*; D013379:Substrate Specificity; D003313:Zea mays / Q000378:metabolism*
D018832:Molecular Chaperones; D011994:Recombinant Proteins; D010444:Peptide Elongation Factor Tu