Information

NpR6012g4 - Biology


Class: Cyanobacteriochrome

Family: Canonical Red/Green

Origin: Nostoc punctiforme

Chromophore(s): PCB

Figure 1. Proposed photostates of the red/green CBCR NpR6012g4. (A) As a working model, we follow a recent report30 that suggested that the phycocyanobilin chromophore is protonated in the red-absorbing 15Z Pr state (left) but deprotonated in the green-absorbing 15E Pg state (right; protonated B-ring tautomer shown)

Absorption spectrum of Npr6012g4 wild type (solid curves) and the beta-Phe mutant (dashed curves) in the 15Z state (red curve) and 15E (green and yellow curves) states.http://larsenlab.ucdavis.edu/@api/deki/files/1584/NpR6012g4.txt

References

  • Femtosecond Photodynamics of the Red/Green Cyanobacteriochrome NpR6012g4 from Nostoc punctiforme: 1. Forward Dynamics, Peter W. Kim, Lucy H. Freer, Nathan C. Rockwell, Shelley S. Martin, J. Clark Lagarias, and Delmar S. Larsen, Biochemistry, 51, 608−618 (2012). pdf
  • Femtosecond Photodynamics of the Red/Green Cyanobacteriochrome NpR6012g4 from Nostoc punctiforme: 2. Reverse Dynamics, Peter W. Larsen, Biochemistry, 51, 619−630 (2012). pdf
  • Second-Chance Initiation Dynamics of the Cyanobacterial Photocycle in the NpR6012 GAF4 Domain of Nostoc Punctiforme, Peter W. Larsen, Journal of the American Chemical Society, 134 (1), 130–133 (2012). pdf
  • Reactive Ground-State Pathways Are Not Ubiquitous in Red/Green Cyanobacteriochromes, Che-Wei Chang, Sean M. Gottlieb, Peter W. Kim, Nathan C. Rockwell, J. Larsen, J. Phys. Chem. B, 117 (38), 11229–11238 (2013). pdf
  • Red/Green Cyanobacteriochromes: Sensors of Color and Power Nathan C. Martin, and J. Clark Lagarias* Department of Molecular and Cellular Biology, University of California, Davis, California 95616, United States Biochemistry, 2012, 51 (48), pp 9667–9677 DOI: 10.1021/bi3013565 Publication Date (Web): November 14, 2012
  • Characterization of Red/Green Cyanobacteriochrome NpR6012g4 by Solution Nuclear Magnetic Resonance Spectroscopy: A Hydrophobic Pocket for the C15-E,anti Chromophore in the Photoproduct, Nathan C. Martin, Sunghyuk Lim, J. Clark Lagarias, and James B. Ames, Biochemistry 2015 54 (24), 3772-3783, DOI: 10.1021/acs.biochem.5b00438 PDF
  • Characterization of Red/Green Cyanobacteriochrome NpR6012g4 by Solution Nuclear Magnetic Resonance Spectroscopy: A Protonated Bilin Ring System in Both Photostates, Nathan C. Ames, Biochemistry 2015 54 (16), 2581-2600, DOI: 10.1021/bi501548t PDF
  • Conservation and Diversity in the Primary Forward Photodynamics of Red/Green Cyanobacteriochromes, Sean M. Kim, Che-Wei Chang, Samuel J. Hanke, Randeep J. Hayer, Nathan C. Clark Lagarias, Delmar S. Larsen, Biochemistry, 54 (4), pp 1028–1042 (2015). PDF
  • Conserved Phenylalanine Residues Are Required for Blue-Shifting of Cyanobacteriochrome Photoproducts, Nathan C. Martin, Alexander G. Gulevich, and J. Clark Lagarias, Biochemistry 2014 53 (19), 3118-3130, DOI: 10.1021/bi500037a PDF

  • NpR3784 is the prototype for a distinctive group of red/green cyanobacteriochromes using alternative Phe residues for photoproduct tuning, Nathan C. Martin, Fei Gan, Donald A. Bryant and J. Clark Lagarias, Photochem. Photobiol. Sci., 2015,14, 258-269, DOI: 10.1039/C4PP00336E PDF

  • Correlating Structural and Photochemical Heterogeneity in Cyanobacteriochrome NpR6012g4, Sunghyuk Lim, Qinhong Yu, Sean M. Gottlieb, Che-Wei Chang, Nathan C. Martin, Dorte Madsen, J. Larsen, and James B. Ames, Proceedings of the National Academy of Sciences of the United States of America, 115 (17) 4387-4392 (2018). pdf


NpR3784 is the prototype for a distinctive group of red/green cyanobacteriochromes using alternative Phe residues for photoproduct tuning

Cyanobacteriochromes (CBCRs) are photosensory proteins found in cyanobacteria and are distantly related to the widespread phytochromes. Whereas plant phytochromes exhibit responses to red and far-red light, CBCRs use the same photoisomerization of a linear tetrapyrrole (bilin) chromophore to respond to a wide range of colors. NpR6012g4 from Nostoc punctiforme and AnPixJ from Anabaena sp. PCC 7120 belong to a large subfamily of red/green CBCRs that exhibit a red-absorbing dark state similar to that of phytochrome but a green-absorbing photoproduct rather than a far-red-absorbing one. In these canonical red/green CBCRs, the photoproduct is blue-shifted relative to the orange absorption observed in the absence of native protein structure. This spectral tuning of the photoproduct requires a conserved Phe residue on the second β strand of the CBCR GAF domain, consistent with a trapped-twist mechanism in which the bilin is sterically constrained in the photoproduct. N. punctiforme also produces NpR3784, a CBCR with a similar red/green photocycle to that of NpR6012g4. NpR3784 lacks both the β2 Phe and other residues characteristic of the canonical red/green CBCRs. In the current work, we identify NpR3784 homologs with red/green photocycles in other cyanobacteria. Spectral tuning in this NpR3784 group is accomplished by a different set of conserved Phe residues, including a characteristic Phe residue on β1. This set of Phe residues cannot be interchanged with the Phe residues found in canonical red/green CBCRs such as NpR6012g4. Our results provide new insights into the flexible protein-chromophore interactions used by CBCRs to generate their remarkable spectral diversity.


NpR6012g4 - Biology

a Department of Molecular and Cell Biology, University of California at Davis, Davis, CA 95616, USA
E-mail: [email protected]

b Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, PA 16802, USA

c Department of Chemistry and Biochemistry, Montana State University, Bozeman, MT 59717, USA

Abstract

Cyanobacteriochromes (CBCRs) are photosensory proteins found in cyanobacteria and are distantly related to the widespread phytochromes. Whereas plant phytochromes exhibit responses to red and far-red light, CBCRs use the same photoisomerization of a linear tetrapyrrole (bilin) chromophore to respond to a wide range of colors. NpR6012g4 from Nostoc punctiforme and AnPixJ from Anabaena sp. PCC 7120 belong to a large subfamily of red/green CBCRs that exhibit a red-absorbing dark state similar to that of phytochrome but a green-absorbing photoproduct rather than a far-red-absorbing one. In these canonical red/green CBCRs, the photoproduct is blue-shifted relative to the orange absorption observed in the absence of native protein structure. This spectral tuning of the photoproduct requires a conserved Phe residue on the second β strand of the CBCR GAF domain, consistent with a trapped-twist mechanism in which the bilin is sterically constrained in the photoproduct. N. punctiforme also produces NpR3784, a CBCR with a similar red/green photocycle to that of NpR6012g4. NpR3784 lacks both the β2 Phe and other residues characteristic of the canonical red/green CBCRs. In the current work, we identify NpR3784 homologs with red/green photocycles in other cyanobacteria. Spectral tuning in this NpR3784 group is accomplished by a different set of conserved Phe residues, including a characteristic Phe residue on β1. This set of Phe residues cannot be interchanged with the Phe residues found in canonical red/green CBCRs such as NpR6012g4. Our results provide new insights into the flexible protein–chromophore interactions used by CBCRs to generate their remarkable spectral diversity.


Auldridge ME, Forest KT (2011) Bacterial phytochromes: more than meets the light. Crit Rev Biochem Mol Biol 46:67–88

Delaglio F, Grzesiek S, Vuister GW, Zhu G, Pfeiffer J, Bax A (1995) NMRPipe: a multidimensional spectral processing system based on UNIX pipes. J Biomol NMR 6:277–293

Gambetta GA, Lagarias JC (2001) Genetic engineering of phytochrome biosynthesis in bacteria. Proc Natl Acad Sci USA 98:10566–10571

Ikeuchi M, Ishizuka T (2008) Cyanobacteriochromes: a new superfamily of tetrapyrrole-binding photoreceptors in cyanobacteria. Photochem Photobiol Sci 7:1159–1167

Ikura M, Kay LE, Bax A (1990) A novel approach for sequential assignment of 1H, 13C, and 15N spectra of proteins: heteronuclear triple-resonance three-dimensional NMR spectroscopy. Application to calmodulin. Biochemistry 29:4659–4667

Kim PW, Freer LH, Rockwell NC, Martin SS, Lagarias JC, Larsen DS (2012) Femtosecond photodynamics of the red/green cyanobacteriochrome NpR6012g4 from Nostoc punctiforme. 1. Forward dynamics. Biochemistry 51:608–618

Narikawa R, Ishizuka T, Muraki N, Shiba T, Kurisu G, Ikeuchi M (2013) Structures of cyanobacteriochromes from phototaxis regulators AnPixJ and TePixJ reveal general and specific photoconversion mechanism. Proc Natl Acad Sci USA 110:918–923

Rockwell NC, Martin SS, Lagarias JC (2012) Red/green cyanobacteriochromes: sensors of color and power. Biochemistry 51:9667–9677

Rockwell NC, Martin SS, Gulevich AG, Lagarias JC (2014) Conserved phenylalanine residues are required for blue-shifting of cyanobacteriochrome photoproducts. Biochemistry 53:3118–3130

Rockwell NC, Martin SS, Lim S, Lagarias JC, Ames JB (2015a) Characterization of red/green cyanobacteriochrome NpR6012g4 by solution nuclear magnetic resonance spectroscopy: a hydrophobic pocket for the C15-E, anti chromophore in the photoproduct. Biochemistry 54:3772–3783

Rockwell NC, Martin SS, Lim S, Lagarias JC, Ames JB (2015b) Characterization of red/green cyanobacteriochrome NpR6012g4 by solution nuclear magnetic resonance spectroscopy: a protonated bilin ring system in both photostates. Biochemistry 54:2581–2600

Wishart DS, Sykes BD, Richards FM (1992) The chemical shift index: a fast and simple method for the assignment of protein secondary structure through NMR spectroscopy. Biochemistry 31:1647–1651


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Title: Evolution-inspired design of multicolored photoswitches from a single cyanobacteriochrome scaffold

Cyanobacteriochromes (CBCRs) are small, bistable linear tetrapyrrole (bilin)-binding light sensors which are typically found as modular components in multidomain cyanobacterial signaling proteins. The CBCR family has been categorized into many lineages that roughly correlate with their spectral diversity, but CBCRs possessing a conserved DXCF motif are found in multiple lineages. DXCF CBCRs typically possess two conserved Cys residues: a first Cys that remains ligated to the bilin chromophore and a second Cys found in the DXCF motif. The second Cys often forms a second thioether linkage, providing a mechanism to sense blue and violet light. DXCF CBCRs have been described with blue/green, blue/orange, blue/teal, and green/teal photocycles, and the molecular basis for some of this spectral diversity has been well established. We here characterize AM1_1499g1, an atypical DXCF CBCR that lacks the second cysteine residue and exhibits an orange/green photocycle. Based on prior studies of CBCR spectral tuning, we have successfully engineered seven AM1_1499g1 variants that exhibit robust yellow/teal, green/teal, blue/teal, orange/yellow, yellow/green, green/green, and blue/green photocycles. The remarkable spectral diversity generated by modification of a single CBCR provides a good template for multiplexing synthetic photobiology systems within the same cellular context, thereby bypassing the time-consuming empirical optimization process neededmore » for multiple probes with different protein scaffolds. « less


NpR6012g4 - Biology

Experimental Data Snapshot

  • Method: SOLUTION NMR
  • Conformers Calculated: 200 
  • Conformers Submitted: 10 
  • Selection Criteria: structures with the lowest energy 

wwPDB Validation   3D Report Full Report

Correlating structural and photochemical heterogeneity in cyanobacteriochrome NpR6012g4.

(2018) Proc Natl Acad Sci U S A 115: 4387-4392

  • PubMed: 29632180  Search on PubMedSearch on PubMed Central
  • DOI: 10.1073/pnas.1720682115
  • Primary Citation of Related Structures:  
    6BHO, 6BHN
  • PubMed Abstract: 

Phytochrome photoreceptors control plant growth, development, and the shade avoidance response that limits crop yield in high-density agricultural plantings. Cyanobacteriochromes (CBCRs) are distantly related photosensory proteins that control cyanobacterial metabolism and behavior in response to light .

Phytochrome photoreceptors control plant growth, development, and the shade avoidance response that limits crop yield in high-density agricultural plantings. Cyanobacteriochromes (CBCRs) are distantly related photosensory proteins that control cyanobacterial metabolism and behavior in response to light. Photoreceptors in both families reversibly photoconvert between two photostates via photoisomerization of linear tetrapyrrole (bilin) chromophores. Spectroscopic and biochemical studies have demonstrated heterogeneity in both photostates, but the structural basis for such heterogeneity remains unclear. We report solution NMR structures for both photostates of the red/green CBCR NpR6012g4 from Nostoc punctiforme In addition to identifying structural changes accompanying photoconversion, these structures reveal structural heterogeneity for residues Trp655 and Asp657 in the red-absorbing NpR6012g4 dark state, yielding two distinct environments for the phycocyanobilin chromophore. We use site-directed mutagenesis and fluorescence and absorbance spectroscopy to assign an orange-absorbing population in the NpR6012g4 dark state to the minority configuration for Asp657. This population does not undergo full, productive photoconversion, as shown by time-resolved spectroscopy and absorption spectroscopy at cryogenic temperature. Our studies thus elucidate the spectral and photochemical consequences of structural heterogeneity in a member of the phytochrome superfamily, insights that should inform efforts to improve photochemical or fluorescence quantum yields in the phytochrome superfamily.


Auldridge ME, Forest KT (2011) Bacterial phytochromes: more than meets the light. Crit Rev Biochem Mol Biol 46:67–88

Campbell EL, Hagen KD, Chen R, Risser DD, Ferreira DP, Meeks JC (2015) Genetic analysis reveals the identity of the photoreceptor for phototaxis in hormogonium filaments of Nostoc punctiforme. J Bacteriol 197:782–791

Enomoto G, Ni-Ni W, Narikawa R, Ikeuchi M (2015) Three cyanobacteriochromes work together to form a light color-sensitive input system for c-di-GMP signaling of cell aggregation. Proc Natl Acad Sci USA 112:8082–8087

Gambetta GA, Lagarias JC (2001) Genetic engineering of phytochrome biosynthesis in bacteria. Proc Natl Acad Sci USA 98:10566–10571

Ikeuchi M, Ishizuka T (2008) Cyanobacteriochromes: a new superfamily of tetrapyrrole-binding photoreceptors in cyanobacteria. Photochem Photobiol Sci 7:1159–1167

Kim PW, Freer LH, Rockwell NC, Martin SS, Lagarias JC, Larsen DS (2012) Femtosecond photodynamics of the red/green cyanobacteriochrome NpR6012g4 from Nostoc punctiforme. 1. Forward dynamics. Biochemistry 51:608–618

Narikawa R, Ishizuka T, Muraki N, Shiba T, Kurisu G, Ikeuchi M (2013) Structures of cyanobacteriochromes from phototaxis regulators AnPixJ and TePixJ reveal general and specific photoconversion mechanism. Proc Natl Acad Sci USA 110:918–923

Rockwell NC, Martin SS, Lagarias JC (2012) Red/green cyanobacteriochromes: sensors of color and power. Biochemistry 51:9667–9677

Rockwell NC, Martin SS, Gulevich AG, Lagarias JC (2014) Conserved phenylalanine residues are required for blue-shifting of cyanobacteriochrome photoproducts. Biochemistry 53:3118–3130

Rockwell NC, Martin SS, Lim S, Lagarias JC, Ames JB (2015a) Characterization of red/green cyanobacteriochrome NpR6012g4 by solution nuclear magnetic resonance spectroscopy: a hydrophobic pocket for the C15-E, anti chromophore in the photoproduct. Biochemistry 54:3772–3783

Rockwell NC, Martin SS, Lim S, Lagarias JC, Ames JB (2015b) Characterization of red/green cyanobacteriochrome NpR6012g4 by solution nuclear magnetic resonance spectroscopy: a protonated bilin ring system in both photostates. Biochemistry 54:2581–2600

Velazquez F, Utesch T, Narikawa R, Ikeuchi M, Mroqinski MA, Gartner W, Hildebrandt P (2013) Photoconversion mechanism of the second GAF domain of cyanobacteriochrome AnPixJ and the cofactor structure of its green-absorbing state. Biochemistry 52:4871–4880

Wishart DS, Sykes BD, Richards FM (1992) The chemical shift index: a fast and simple method for the assignment of protein secondary structure through NMR spectroscopy. Biochemistry 31:1647–1651


Cyanobacteriochromes: photoreceptors covering the entire UV-to-visible spectrum

Combination of four color-tuning mechanisms results in diverse spectral properties.

Structural information uncovers general and specific photoconversion mechanisms.

Protein engineering studies for optogenetic tools are now proceeding.

Cyanobacteriochrome photoreceptors are linear tetrapyrrole-binding photoreceptors that are distantly related to the canonical phytochrome photoreceptors. The chromophore-binding region of the cyanobacteriochromes consists of only a cGMP-phosphodiesterase/adenylate cyclase/FhlA (GAF) domain, while that of the phytochromes consists of three domains, including the GAF domain. Most of the canonical phytochromes homogenously show red/far-red reversible photoconversion. Conversely, the cyanobacteriochrome photoreceptors are highly diverse in the colors of light they sense. Since the discovery of the first cyanobacteriochrome photoreceptor around 15 years ago, physiological, biochemical, and biophysical studies on cyanobacteriochromes have been extensively performed to date. In this review, we focus on color-tuning mechanisms of diverse cyanobacteriochromes.


NpR3784 is the prototype for a distinctive group of red/green cyanobacteriochromes using alternative Phe residues for photoproduct tuning

Cyanobacteriochromes (CBCRs) are photosensory proteins found in cyanobacteria and are distantly related to the widespread phytochromes. Whereas plant phytochromes exhibit responses to red and far-red light, CBCRs use the same photoisomerization of a linear tetrapyrrole (bilin) chromophore to respond to a wide range of colors. NpR6012g4 from Nostoc punctiforme and AnPixJ from Anabaena sp. PCC 7120 belong to a large subfamily of red/green CBCRs that exhibit a red-absorbing dark state similar to that of phytochrome but a green-absorbing photoproduct rather than a far-red-absorbing one. In these canonical red/green CBCRs, the photoproduct is blue-shifted relative to the orange absorption observed in the absence of native protein structure. This spectral tuning of the photoproduct requires a conserved Phe residue on the second &beta strand of the CBCR GAF domain, consistent with a trapped-twist mechanism in which the bilin is sterically constrained in the photoproduct. N. punctiforme also produces NpR3784, a CBCR with a similar red/green photocycle to that of NpR6012g4. NpR3784 lacks both the &beta2 Phe and other residues characteristic of the canonical red/green CBCRs. In the current work, we identify NpR3784 homologs with red/green photocycles in other cyanobacteria. Spectral tuning in this NpR3784 group is accomplished by a different set of conserved Phe residues, including a characteristic Phe residue on &beta1. This set of Phe residues cannot be interchanged with the Phe residues found in canonical red/green CBCRs such as NpR6012g4. Our results provide new insights into the flexible protein&ndashchromophore interactions used by CBCRs to generate their remarkable spectral diversity.

Journal

Photochemical & Photobiological Sciences &ndash Royal Society of Chemistry