Resonance Raman Spectroscopy Reveals the Origin of an Intermediate Wavelength Form in Photoactive Yellow Protein
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- 作者：Samir F. El-Mashtoly ; Masashi Unno ; Masato Kumauchi ; Norio Hamada ; Kimiyo Fujiwara ; Jun Sasaki ; Yasushi Imamoto ; Mikio Kataoka ; Fumio Tokunaga ; and Seigo Yamauchi
- 刊名：Biochemistry
- 出版年：2004
- 出版时间：March 2, 2004
- 年：2004
- 卷：43
- 期：8
- 页码：2279 - 2287
- 全文大小：182K
- 年卷期：v.43,no.8(March 2, 2004)
- ISSN：1520-4995
文摘
Photoactive yellow protein (PYP) is a bacterial blue light receptor containing a 4-hydroxycinnamyl chromophore, and its absorption maximum is 446 nm. In a dark state, the hydroxyl group of thechromophore is deprotonated and forms hydrogen bonds with Tyr42 and Glu46. Either removal of ahydrogen bond with Tyr42 or addition of chaotropes such as thiocyanate produces a blue-shifted speciescalled an intermediate wavelength form, in which absorption maximum ranges from 355 to 400 nm. Toexamine the structural origin of the intermediate wavelength form, we have performed resonance Ramaninvestigations of wild-type PYP and some mutants (Tyr42 
f"> Ala, Tyr42 f"> Phe, Glu46 f"> Gln, andThr50 f"> Val) in the presence or absence of potassium thiocyanate. These studies show that the chromophoreof the intermediate wavelength form is protonated, implying an increase in a pKa of the chromophore.Hence, the removal of the hydrogen bond between Tyr42 and chromophore or partial protein denaturationin the presence of thiocyanate results in a spectral blue-shift. Quantum chemical calculations based ondensity functional theory further support the idea that the pKa of the chromophore is increased by removinga hydrogen bond or by increasing the dielectric constant in the vicinity of the chromophore.
f"> Ala, Tyr42 f"> Phe, Glu46 f"> Gln, andThr50 f"> Val) in the presence or absence of potassium thiocyanate. These studies show that the chromophoreof the intermediate wavelength form is protonated, implying an increase in a pKa of the chromophore.Hence, the removal of the hydrogen bond between Tyr42 and chromophore or partial protein denaturationin the presence of thiocyanate results in a spectral blue-shift. Quantum chemical calculations based ondensity functional theory further support the idea that the pKa of the chromophore is increased by removinga hydrogen bond or by increasing the dielectric constant in the vicinity of the chromophore.