Expression, purification and characterization of a novel bZIP protein from sugarcane

Abstract

The basic leucine zipper (bZIP) proteins form a large family of transcriptional factors in plants and other eukaryotes. Plant bZIP transcriptional factors are divided into subfamilies and are involved in regulating a large range of physiological processes, from plant development to responses to biotic and abiotic stimuli. In this work, we cloned a novel bZIP of sugarcane into the pET3c vector and expressed the recombinant SCbZIP1 (66-170) protein in Escherichia coli BL21 (DE3) plysS. The recombinant protein was purified by heat-treatment and reversed phase chromatography. Northern blot analysis showed that SCbZIP1 was expressed early in development on day 4, but was not induced by abscisic acid (ABA) or exposure to cold. The identity of the recombinant protein was confirmed by mass spectrometry and CD spectroscopy showed an alpha-helical content of 33%. Electrophoretic mobility assays showed that SCbZIP1 (66-170) bound strongly to G-box, Hex and C-box DNA motifs. SCbZIP1 (66-170) was phosphorylated in vitro by a series of protein kinases and its DNA-binding affinity was strongly decreased after phosphorylation by CKII. SCbZIP1 (66-170) also underwent homo- and heterodimerization with truncated forms of the bZIP transcription factor Opaque 2 from Coix.