PURIFICATION AND CHARACTERIZATION OF AN AUXIN-BINDING PROTEIN FROM ARABIDOPSIS-THALIANA EXPRESSED IN BACULOVIRUS-INFECTED INSECT CELLS

The auxin-binding protein At-ERabpl is of very low abundance in Arabid opsis thaliana; it hinders any study at the protein level as it is dif ficult to collect large amounts from the plant. We therefore chose to express At-ERabpl in baculovirus-infected insect cells. Recombinant ba culoviruses were selected in yeast according to Patel et al. (Nucleic Acids Res. 20, 97-104, 1991). The recombinant protein was purified to homogeneity by a simple procedure involving an affinity step on a succ inyl-concanavalin A column, Labeling with the photoactive auxin 5-N-3- [7-H-3]indole-3-acetic acid demonstrated that the baculovirus-expresse d protein belongs to the auxin-binding protein family as deduced from its cDNA homology to a gene previously characterized in maize. The mat ure polypeptide migrates on SDS-PAGE with an apparent molecular mass o f about 23 kDa, its NH2-leader sequence is properly processed, and it bears a high-mannose-type sugar moiety, All results are in agreement w ith information derived from the cDNA analysis. The possible role of a functional dimerization is also discussed. (C) 1995 Academic Press, I nc.