Target sequence recognition by a light-activatable bZIP factor, Photozipper.

Photozipper (PZ) is a light-activatable basic leucine zipper (bZIP) protein composed of a bZIP domain and a light-oxygen-voltage-sensing domain of aureochrome-1. Blue light induces dimerization and subsequently increases the affinity of PZ for the target DNA sequence. We prepared site-directed PZ mutants in which Asn131 (N131) in the basic region was substituted with Ala and Gln. N131 mutants showed spectroscopic and dimerization properties almost identical to those of wild-type PZ and an increase in helical content in the presence of the target sequence. Quantitative analyses by an electrophoretic mobility shift assay and quartz crystal microbalance (QCM) measurements demonstrated that the half-maximal effective concentrations of N131 mutants to bind to the target sequence were significantly higher than those of PZ. QCM data also revealed that N131 substitutions accelerated the dissociation without affecting the association, suggesting that a base-specific interaction of N131 occurred after the association between PZ and DNA. Activation of PZ by illumination decreased both the standard errors and the unstable period of QCM data. Optical control of transcription factors will provide new knowledge of the recognition of the target sequence.