Natural diversity provides a broad spectrum of cyanobacteriochrome-based diguanylate cyclases

Blain-Hartung, Matthew; Rockwell, Nathan C.; Lagarias, J. Clark

doi:10.1093/plphys/kiab240

Cited by 14 publications

(12 citation statements)

References 80 publications

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“…Whereas the PAS-GAF core is rigidly conserved, orientation of the PHY dimer interface and the central helical spine remains malleable. Potentially, this could also be one explanation for why PadCs tend to allow higher dynamic ranges than many PHY-less cyanobacteriochrome-based diguanylate cyclases [ 49 ]. However, tight interaction of the central helical spine is not necessarily a functional prerequisite for other bacteriophytochromes, as observed for the phytochrome-linked histidine kinase/phosphatase of Deinococcus radiodurans with its relatively loose PHY domain interaction [ 50 ] or the Xanthomonas campestris BphP where breaks of the helical spine affect the quaternary assembly [ 46 ].…”

Section: Discussionmentioning

confidence: 99%

Characterisation of sequence–structure–function space in sensor–effector integrators of phytochrome-regulated diguanylate cyclases

Böhm

Gourinchas

Zweytick

et al. 2022

Photochem Photobiol Sci

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Understanding the relationship between protein sequence, structure and function is one of the fundamental challenges in biochemistry. A direct correlation, however, is often not trivial since protein dynamics also play an important functional role—especially in signal transduction processes. In a subfamily of bacterial light sensors, phytochrome-activated diguanylate cyclases (PadCs), a characteristic coiled-coil linker element connects photoreceptor and output module, playing an essential role in signal integration. Combining phylogenetic analyses with biochemical characterisations, we were able to show that length and composition of this linker determine sensor–effector function and as such are under considerable evolutionary pressure. The linker length, together with the upstream PHY-specific domain, influences the dynamic range of effector activation and can even cause light-induced enzyme inhibition. We demonstrate phylogenetic clustering according to linker length, and the development of new linker lengths as well as new protein function within linker families. The biochemical characterisation of PadC homologs revealed that the functional coupling of PHY dimer interface and linker element defines signal integration and regulation of output functionality. A small subfamily of PadCs, characterised by a linker length breaking the coiled-coil pattern, shows a markedly different behaviour from other homologs. The effect of the central helical spine on PadC function highlights its essential role in signal integration as well as direct regulation of diguanylate cyclase activity. Appreciation of sensor–effector linkers as integrator elements and their coevolution with sensory modules is a further step towards the use of functionally diverse homologs as building blocks for rationally designed optogenetic tools. Graphical abstract

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Section: Discussionmentioning

confidence: 99%

Characterisation of sequence–structure–function space in sensor–effector integrators of phytochrome-regulated diguanylate cyclases

Böhm

Gourinchas

Zweytick

et al. 2022

Photochem Photobiol Sci

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“…In PVB-binding GAF domains, PCB is initially incorporated into the CBCR GAF domain, followed by the isomerization of PCB to PVB [6,9]. Among the PCB-binding CBCR GAF domains, many GAF domains form a huge expanded red/green (XRG) lineage [7,[10][11][12][13][14]. Typical XRG GAF domains covalently bind PCB and show red/green reversible photoconversion, but many atypical ones have been identified to show violet/blue, violet/orange, red/blue, or green/blue photoconversion.…”

Section: Introductionmentioning

confidence: 99%

“…Most CBCR GAF domains possess highly conserved Cys residue (canonical Cys) for stable chromophore ligation [1]. In addition, many CBCR GAF domains have a second conserved Cys residue (second Cys) [4][5][6]10,12,14,[19][20][21][22][23][24][25][26]. The typical role of the second Cys residue is reversible or stable attachment to C10 of the chromophore, which vastly shortens the p-conjugated system of the chromophore, leading to UV-to-blue absorption.…”

Section: Introductionmentioning

confidence: 99%

“…Notably, the second Cys residues-from structurally different positions-have been reported to ligate to C10 of the chromophore. Four distinct structural positions have been identified for the second Cys residue [10,12,14,19]. Of these, the second Cys residue within the highly conserved DXCF motif has an additional function in that it is crucial for isomerization from PCB to PVB upon chromophore incorporation [6,9,21].…”

Section: Introductionmentioning

confidence: 99%

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Introduction of reversible cysteine ligation ability to the biliverdin‐binding cyanobacteriochrome photoreceptor

Suzuki,

Yoshimura,

Hoshino

et al. 2023

The FEBS Journal

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Cyanobacteriochrome (CBCR) photoreceptors are distantly related to the canonical red/far‐red reversible phytochrome photoreceptors. In the case of the CBCRs, only the GAF domain is required for chromophore incorporation and photoconversion. The GAF domains of CBCR are highly diversified into many lineages to sense various colors of light. These CBCR GAF domains are divided into two types: those possessing only the canonical Cys residue and those with both canonical and second Cys residues. The canonical Cys residue stably ligates to the chromophore in both cases. The second Cys residue mostly shows reversible adduct formation with the chromophore during photoconversion for spectral tuning. In this study, we focused on the CBCR GAF domain AnPixJg2_BV4, which possesses only the canonical Cys residue. AnPixJg2_BV4 covalently ligates to the biliverdin (BV) chromophore and shows far‐red/orange reversible photoconversion. Because BV is a mammalian intrinsic chromophore, BV‐binding molecules are advantageous for in vivo optogenetic and bioimaging tool development. To obtain a better developmental platform molecule, we performed site‐saturation random mutagenesis and serendipitously obtained a unique variant molecule that showed far‐red/blue reversible photoconversion, in which the Cys residue was introduced near the chromophore. This introduced Cys residue functioned as the second Cys residue that reversibly ligated with the chromophore. Because the position of the introduced Cys residue is distinct from the known second Cys residues, the variant molecule obtained in this study would expand our knowledge about the spectral tuning mechanism of CBCRs and contribute to tool development.

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“…Such studies can also provide new insights for protein engineering efforts. There is growing interest in developing red-responsive tools based on CBCRs [46,[99][100][101][102], so the mechanisms that determine whether a given domain undergoes photoconversion or exhibits other de-excitation mechanisms can also inform the choice of initial mutations for development of a fluorescent or photoactive reporter which could then be subjected to further optimization.…”

Section: Introductionmentioning

confidence: 99%

Protein–chromophore interactions controlling photoisomerization in red/green cyanobacteriochromes

Rockwell

Moreno

Martin

et al. 2022

Photochem Photobiol Sci

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Photoreceptors in the phytochrome superfamily use 15,16-photoisomerization of a linear tetrapyrrole (bilin) chromophore to photoconvert between two states with distinct spectral and biochemical properties. Canonical phytochromes include master regulators of plant growth and development in which light signals trigger interconversion between a red-absorbing 15Z dark-adapted state and a metastable, far-red-absorbing 15E photoproduct state. Distantly related cyanobacteriochromes (CBCRs) carry out a diverse range of photoregulatory functions in cyanobacteria and exhibit considerable spectral diversity. One widespread CBCR subfamily typically exhibits a red-absorbing 15Z dark-adapted state similar to that of phytochrome that gives rise to a distinct green-absorbing 15E photoproduct. This red/green CBCR subfamily also includes red-inactive examples that fail to undergo photoconversion, providing an opportunity to study protein–chromophore interactions that either promote photoisomerization or block it. In this work, we identified a conserved lineage of red-inactive CBCRs. This enabled us to identify three substitutions sufficient to block photoisomerization in photoactive red/green CBCRs. The resulting red-inactive variants faithfully replicated the fluorescence and circular dichroism properties of naturally occurring examples. Converse substitutions restored photoconversion in naturally red-inactive CBCRs. This work thus identifies protein–chromophore interactions that control the fate of the excited-state population in red/green cyanobacteriochromes.

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Natural diversity provides a broad spectrum of cyanobacteriochrome-based diguanylate cyclases

Cited by 14 publications

References 80 publications

Characterisation of sequence–structure–function space in sensor–effector integrators of phytochrome-regulated diguanylate cyclases

Characterisation of sequence–structure–function space in sensor–effector integrators of phytochrome-regulated diguanylate cyclases

Introduction of reversible cysteine ligation ability to the biliverdin‐binding cyanobacteriochrome photoreceptor

Protein–chromophore interactions controlling photoisomerization in red/green cyanobacteriochromes

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