Evidence for <i>trans‐cis</i> isomerization of the <i>p</i>‐coumaric acid chromophore as the photochemical basis of the photocycle of photoactive yellow protein

Kort, Remco; Vonk, Heleen S.; Xu, Xiufang; Hoff, Wouter D.; Crielaard, Wim; Hellingwerf, Klaas J.

doi:10.1016/0014-5793(96)00149-4

Cited by 212 publications

(192 citation statements)

References 26 publications

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“…The emerging mechanism of receptor activation in PYP contains the following steps: (i) initial chromophore photoisomerization (32), which triggers (ii) active site proton transfer (33) that causes (iii) an electrostatically triggered protein quake (12), resulting in structural disruption of the PAS domain core, which is relayed (iv) to the N-terminal helices (13,14) by the side chain of Asn-43 (34). Future studies are needed to reveal how partial PAS domain unfolding contributes to signaling and whether this mechanism operates more widely in the PAS domain family.…”

Section: Discussionmentioning

confidence: 99%

Single-molecule detection of structural changes during Per-Arnt-Sim (PAS) domain activation

Zhao

Lee²,

Nome

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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The Per-Arnt-Sim (PAS) domain is a ubiquitous protein module with a common three-dimensional fold involved in a wide range of regulatory and sensory functions in all domains of life. The activation of these functions is thought to involve partial unfolding of N-or C-terminal helices attached to the PAS domain. Here we use atomic force microscopy to probe receptor activation in single molecules of photoactive yellow protein (PYP), a prototype of the PAS domain family. Mechanical unfolding of Cys-linked PYP multimers in the presence and absence of illumination reveals that, in contrast to previous studies, the PAS domain itself is extended by Ϸ3 nm (at the 10-pN detection limit of the measurement) and destabilized by Ϸ30% in the light-activated state of PYP. Comparative measurements and steered molecular dynamics simulations of two double-Cys PYP mutants that probe different regions of the PAS domain quantify the anisotropy in stability and changes in local structure, thereby demonstrating the partial unfolding of their PAS domain upon activation. These results establish a generally applicable single-molecule approach for mapping functional conformational changes to selected regions of a protein. In addition, the results have profound implications for the molecular mechanism of PAS domain activation and indicate that stimulusinduced partial protein unfolding can be used as a signaling mechanism.atomic force spectroscopy ͉ photoactive yellow protein ͉ receptor activation B iological signaling starts with the activation of receptor proteins, in which stimuli trigger protein conformational changes that relay a signal to an associated signal transduction chain. A central question concerns the nature of the structural changes that activate a receptor protein. The mechanism of signaling by PerArnt-Sim (PAS) domain-containing proteins is of considerable interest, because these structural domains occur in many signaling proteins (1). In addition, incorrect signaling by PAS domains is associated with a range of diseases. Important examples are mutations in the human ERG potassium channel, which causes cardiac arrhythmias (2), and the involvement of hypoxia-inducible factors in myocardial and cerebral ischemia (3).We investigate the photoinduced structural changes in the bacterial blue light receptor, photoactive yellow protein (PYP), a prototype for PAS domains (4). As shown in Fig. 1, PYP consists of two N-terminal ␣-helices (residues 1-25), followed by a typical PAS domain (1, 5) fold (residues 26-125) that contains a central six-stranded ␤-sheet flanked by ␣-helices (6). The protein exhibits photochemistry based on its p-coumaric acid chromophore (7). Blue light converts the initial pG state of PYP into the photoactivated pB state in milliseconds, whereas pB decays back to pG in Ϸ0.5 s (8). The pB state is believed to trigger negative phototaxis in purple bacteria (9).The structural changes that occur upon pB formation involve partial unfolding of the receptor, indicating partial protein unfolding as a novel signaling m...

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Section: Discussionmentioning

confidence: 99%

Single-molecule detection of structural changes during Per-Arnt-Sim (PAS) domain activation

Zhao

Lee²,

Nome

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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“…4. On absorption of a photon, the chromophore of PYP isomerizes to the cis form (15). There are two possible models for the isomerization of the chromophore: one is that the phenol part rotates, and the other is that the ester part rotates.…”

Section: Discussionmentioning

confidence: 99%

“…On photoconversion from PYP to PYP M , the proton is transferred from Glu-46 to the chromophore. The chromophore of PYP is in the trans form (7,15) whereas that of PYP M is in the cis form (15).…”

Section: Fig 4 Schematic Drawing Of Proton Movement In Pypmentioning

confidence: 99%

“…Despite the striking differences in the chromophores, protein moieties, and linkages, the chromophores of both retinal proteins and PYP turn neutral in their near-UV intermediates, which are thought to be formed as the result of large conformational changes (17,28). In addition, cis/trans isomerization of the chromophore is involved in both systems (15). These are the common events for the protein conformational changes among the photoreceptor proteins and would be the most efficient mechanism.…”

Section: Internal Proton Transfer During Pyp Photocycle 12907mentioning

confidence: 99%

“…Crystallography at 1.4-Å resolution (7) and resonance Raman spectroscopy (14) have shown that the phenolic oxygen of the chromophore of PYP is deprotonated in the dark state. The photocycle is initiated by photon absorption, which involves isomerization of the ethylenic bond of the chromophore (15). During the photocycle, observed proton uptake and release correspond with the formation and decay of PYP M (16).…”

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Evidence for Proton Transfer from Glu-46 to the Chromophore during the Photocycle of Photoactive Yellow Protein

Imamoto

Mihara

Hisatomi

et al. 1997

Journal of Biological Chemistry

125

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Photoactive yellow protein (PYP) belongs to the novel group of eubacterial photoreceptor proteins. To fully understand its light signal transduction mechanisms, elucidation of the intramolecular pathway of the internal proton is indispensable because it closely correlates with the changes in the hydrogen-bonding network, which is likely to induce the conformational changes. For this purpose, the vibrational modes of PYP and its photoproduct were studied by Fourier transform infrared spectroscopy at ؊40°C. The vibrational modes characteristic for the anionic p-coumaryl chromophore (Kim, M., Mathies, R. A., Hoff, W. D., and Hellingwerf, K. J. (1995) Biochemistry 34, 12669 -12672) were observed at 1482, 1437, and 1163 cm ؊1 for PYP. However, the bands corresponding to these modes were not observed for PYP M , the blue-shifted intermediate, but the 1175 cm ؊1 band characteristic of the neutral p-coumaryl chromophore was observed, indicating that the phenolic oxygen of the chromophore is protonated in PYP M . A 1736 cm ؊1 band was observed for PYP, but the corresponding band for PYP M was not. Because it disappeared in the Glu-46 3 Gln mutant of PYP, this band was assigned to the C‫؍‬O stretching mode of the COOH group of Glu-46. These results strongly suggest that the proton at Glu-46 is transferred to the chromophore during the photoconversion from PYP to PYP M .Photoactive yellow protein (PYP) 1 ( max ϭ 446 nm) (1) is proposed to be a photoreceptor protein for the negative phototaxis observed in the purple phototrophic bacterium, Ectothiorhodospira halophila (2). PYP belongs to the novel group of photoreceptor proteins (3, 4) whose structures are quite different from those of the other photoreceptor proteins studied so far. Namely, the protein moiety of PYP has an ␣/␤ fold structure (5) composed of 125 amino acids (6, 7). The chromophore is a p-coumaric acid (7-9) bound to a cysteine residue via a thioester bond.PYP absorbs a photon and enters the photocycle. We have analyzed the photocycle of PYP in detail by low temperature spectroscopy and identified several intermediates (10). Irradiation of PYP at Ϫ190°C yields PYP B ( max ϭ 489 nm) and PYP H ( max ϭ 442 nm), which are thermally converted to PYP L ( max ϭ 456 nm) through PYP BL ( max ϭ 400 nm) and PYP HL ( max ϭ 447 nm), respectively. The two pathways beginning with PYP B and PYP H join at PYP L and revert to PYP. Flash photolysis at ambient temperature identified two intermediates, pR ( max ϭ 465 nm) and pB ( max ϭ 355 nm) (11,12). pR is formed within 10 ns after flash excitation. It decays to pB over a submillisecond time scale and reverts to PYP within 1 s. It has been demonstrated that pR is the same species as PYP L (10) (In this paper, pR and pB are called PYP L and PYP M , respectively, to avoid confusion) and that PYP L is accumulated by irradiation of PYP at Ϫ80°C (10, 13). However, the precursors of PYP L have not been discovered by flash photolysis at room temperature.Recent studies have clarified some details of the events that take place during t...

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Polycephalin B und C: ungewöhnliche Tetramsäuren aus Plasmodien des SchleimpilzesPhysarum polycephalum (Myxomycetes)

Nowak¹,

Steffan²

1998

Angewandte Chemie

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Evidence for trans‐cis isomerization of the p‐coumaric acid chromophore as the photochemical basis of the photocycle of photoactive yellow protein

Cited by 212 publications

References 26 publications

Single-molecule detection of structural changes during Per-Arnt-Sim (PAS) domain activation

Single-molecule detection of structural changes during Per-Arnt-Sim (PAS) domain activation

Evidence for Proton Transfer from Glu-46 to the Chromophore during the Photocycle of Photoactive Yellow Protein

Polycephalin B und C: ungewöhnliche Tetramsäuren aus Plasmodien des SchleimpilzesPhysarum polycephalum (Myxomycetes)

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