Tsuji, A. | Search bioRxiv

bioRxiv2023

Structural basis of EHEP-mediated offense against phlorotannin-induced defense from brown algae to protect akuBGL activity

The defensive-offensive associations between algae and herbivores determine marine ecology. Brown algae utilize phlorotannin as their chemical defense against the predator Aplysia kurodai, which uses {beta}-glucosidase (akuBGL) to digest the laminarin in algae to glucose. Moreover, A. kurodai employs Eisenia hydrolysis-enhancing protein (EHEP) as an offense to protect akuBGL activity from phlorotannin inhibition by precipitating phlorotannin. To underpin the molecular mechanism of this digestive-defensive-offensive system, we determined the structures of apo and tannic-acid (TNA, a phlorotannin-analog) bound form of EHEP, as well as apo akuBGL. EHEP consisted of three peritrophin-A domains formed in a triangle and bound TNA in the center without significant conformational changes. Structural comparison between EHEP and EHEP-TNA led us to find that EHEP can be resolubilized from phlorotannin-precipitation at an alkaline pH, which reflects a requirement in the digestive tract. akuBGL contained two GH1 domains, only one of which conserved the active site. Combining docking analysis, we propose the mechanisms by which phlorotannin inhibits akuBGL by occupying the substrate-binding pocket, and EHEP protects akuBGL against the inhibition by binding with phlorotannin to free the akuBGL pocket.

biochemistry↗

bioRxiv2022

Dimerization processes for light-regulated transcription factor Photozipper visualized by high-speed atomic force microscopy

Dimerization is critical for transcription factors (TFs) to bind DNA and regulate a wide variety of cellular functions; however, the single-molecular mechanisms remain to be completely elucidated. Here, we used high-speed atomic force microscopy (HS-AFM) to observe the dimerization process for a photoresponsive TF Photozipper (PZ), which consists of light-oxygen-voltage-sensing (LOV) and basic-region-leucine-zipper (bZIP) domain. HS-AFM visualized not only the oligomeric states of PZ molecules forming monomers and dimers under controlled dark-light conditions but also the domain structures within each molecule. Successive AFM movies captured the dimerization process for an individual PZ molecule and the monomer-dimer reversible transition during dark-light cycling. The high-resolution AFM images of the domain structures in PZ molecules demonstrated that the bZIP domain entangled under dark conditions was loosened owing to light illumination and fluctuated around the LOV domain. These observations revealed the role of the bZIP domain in the dimerization processes of a TF.

biophysics↗