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Elucidating Ultrafast Multiphasic Dynamics in the Photoisomerization of Cyanobacteriochrome.

ABSTRACT: Understanding photoisomerization dynamics in cyanobacteriochromes is important to the development of optical agents in near-infrared biological imaging and optogenetics. Here, by integrating femtosecond spectroscopy and site-directed mutagenesis, we investigate the photoinduced Pr-state isomerization dynamics and mechanism of a unique red/green cyanobacteriochrome from Leptolyngbya sp. JSC-1. We observed multiphasic dynamics in the Pr state, a widespread phenomenon for photoreceptors in the phytochrome superfamily, and revealed their origins; the initial dynamics over a few to tens and hundreds of picoseconds arises from the local active-site relaxations followed by the slow double-bond isomerization in several hundreds of picoseconds. Such continuous active-site evolution results in a unique spectral tuning effect that favors the blue-side emission and suppresses the red-side emission. We also observed the faster dynamics in both relaxation and isomerization with critical mutants at the active site that render a looser active site. These results clearly distinguish the multiphasic dynamics between relaxation and isomerization and reveal a novel molecular mechanism for better biological applications.

SUBMITTER: Wang D

PROVIDER: S-EPMC8172094 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature

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Elucidating Ultrafast Multiphasic Dynamics in the Photoisomerization of Cyanobacteriochrome.

Wang Dihao D Li Xiankun X Wang Lijuan L Yang Xiaojing X Zhong Dongping D

The journal of physical chemistry letters 20201001 20

Understanding photoisomerization dynamics in cyanobacteriochromes is important to the development of optical agents in near-infrared biological imaging and optogenetics. Here, by integrating femtosecond spectroscopy and site-directed mutagenesis, we investigate the photoinduced Pr-state isomerization dynamics and mechanism of a unique red/green cyanobacteriochrome from <i>Leptolyngbya</i> sp. JSC-1. We observed multiphasic dynamics in the Pr state, a widespread phenomenon for photoreceptors in t ...[more]

PMID: 32940473

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Project description:Cyanobacteriochromes (CBCRs) are promising optogenetic tools for their diverse absorption properties with a single compact cofactor-binding domain. We previously uncovered the ultrafast reversible photoswitching dynamics of a red/green photoreceptor AnPixJg2, which binds phycocyanobilin (PCB) that is unavailable in mammalian cells. Biliverdin (BV) is a mammalian cofactor with a similar structure to PCB but exhibits redder absorption. To improve the AnPixJg2 feasibility in mammalian applications, AnPixJg2_BV4 with only four mutations has been engineered to incorporate BV. Herein, we implemented femtosecond transient absorption (fs-TA) and ground state femtosecond stimulated Raman spectroscopy (GS-FSRS) to uncover transient electronic dynamics on molecular time scales and key structural motions responsible for the photoconversion of AnPixJg2_BV4 with PCB (Bpcb) and BV (Bbv) cofactors in comparison with the parent AnPixJg2 (Apcb). Bpcb adopts the same photoconversion scheme as Apcb, while BV4 mutations create a less bulky environment around the cofactor D ring that promotes a faster twist. The engineered Bbv employs a reversible clockwise/counterclockwise photoswitching that requires a two-step twist on ~5 and 35 picosecond (ps) time scales. The primary forward Pfr → Po transition displays equal amplitude weights between the two processes before reaching a conical intersection. In contrast, the primary reverse Po → Pfr transition shows a 2:1 weight ratio of the ~35 ps over 5 ps component, implying notable changes to the D-ring-twisting pathway. Moreover, we performed pre-resonance GS-FSRS and quantum calculations to identify the Bbv vibrational marker bands at ~659,797, and 1225 cm-1. These modes reveal a stronger H-bonding network around the BV cofactor A ring with BV4 mutations, corroborating the D-ring-dominant reversible photoswitching pathway in the excited state. Implementation of BV4 mutations in other PCB-binding GAF domains like AnPixJg4, AM1_1870g3, and NpF2164g5 could promote similar efficient reversible photoswitching for more directional bioimaging and optogenetic applications, and inspire other bioengineering advances.

Dataset Information

Elucidating Ultrafast Multiphasic Dynamics in the Photoisomerization of Cyanobacteriochrome.

Publications

Elucidating Ultrafast Multiphasic Dynamics in the Photoisomerization of Cyanobacteriochrome.

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