Abstract

Cyanobacteriochromes (CBCRs) are distantly related to the red/far-red responsive phytochromes. Red/green-type CBCRs are widely distributed among various cyanobacteria. The red/green-type CBCRs covalently bind phycocyanobilin (PCB) and show red/green reversible photoconversion. Recent studies revealed that some red/green-type CBCRs from chlorophyll d-bearing cyanobacterium Acaryochloris marina covalently bind not only PCB but also biliverdin (BV). The BV-binding CBCRs show far-red/orange reversible photoconversion. Here, we identified another CBCR (AM1_C0023g2) from A. marina that also covalently binds not only PCB but also BV with high binding efficiencies, although BV chromophore is unstable in the presence of urea. Replacement of Ser334 with Gly resulted in significant improvement in the yield of the BV-binding holoprotein, thereby ensuring that the mutant protein is a fine platform for future development of optogenetic switches. We also succeeded in detecting near-infrared fluorescence from mammalian cells harboring PCB-binding AM1_C0023g2 whose fluorescence quantum yield is 3.0%. Here the PCB-binding holoprotein is shown as a platform for future development of fluorescent probes.

Highlights

  • Phytochromes and cyanobacteriochromes (CBCRs) are photoreceptors that form a large superfamily with a linear tetrapyrrole-binding GAF domain (Ikeuchi and Ishizuka, 2008; Anders and Essen, 2015)

  • Some CBCRs are known to be involved in regulation of light acclimation processes such as phototaxis (Yoshihara et al, 2000; Narikawa et al, 2011; Song et al, 2011; Savakis et al, 2012; Campbell et al, 2015), chromatic acclimation (Kehoe and Grossman, 1996; Hirose et al, 2010) and light-dependent cell aggregation (Enomoto et al, 2014, 2015)

  • GAF domain of CBCRs is enough for chromophore ligation and photoconversion, additional PAS (Per/Arnt/Sim) and PHY

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Summary

INTRODUCTION

Phytochromes and cyanobacteriochromes (CBCRs) are photoreceptors that form a large superfamily with a linear tetrapyrrole-binding GAF (cGMP-phosphodiesterase/adenylate cyclase/FhlA) domain (Ikeuchi and Ishizuka, 2008; Anders and Essen, 2015). PVBbinding CBCRs sense relatively shorter wavelength light covering ultraviolet-to-green region (Yoshihara et al, 2004; Ishizuka et al, 2006; Rockwell et al, 2008, 2012a,b; Narikawa et al, 2011; Song et al, 2011; Enomoto et al, 2012; Ma et al, 2012; Cho et al, 2015), whereas PCB-binding CBCRs sense longer wavelength light covering ultraviolet-to-red region (Hirose et al, 2008, 2013; Narikawa et al, 2008a,b, 2014; Rockwell et al, 2011, 2012c; Chen et al, 2012). BV is present in most organisms including mammals and far-red light can penetrate into deep tissues, with a potential as optogenetic and bioimaging tools (Ziegler and Möglich, 2015) We report another GAF domain (second GAF domain of AM1_C0023 called AM1_C0023g2) from A. marina that covalently binds PCB and BV. We detected near-infrared fluorescence from mammalian cells harboring AM1_C0023g2-PCB whose fluorescence quantum yield is 3.0%

MATERIALS AND METHODS
RESULTS AND DISCUSSION
Evaluation of Chromophore Binding Efficiencies
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