Title
Binding, tuning and mechanical function of the 4-hydroxy-cinnamic acid chromophore in photoactive yellow protein
Author
van der Horst, M.A.
Arents, J.C.
Kort, R.
Hellingwerf, K.J.
TNO Kwaliteit van Leven
Publication year
2007
Abstract
The bacterial photoreceptor protein photoactive yellow protein (PYP) covalently binds the chromophore 4-hydroxy coumaric acid, tuning (spectral) characteristics of this cofactor. Here, we study this binding and tuning using a combination of pointmutations and chromophore analogs. In all photosensor proteins studied to date the covalent linkage of the chromophore to the apoprotein is dispensable for light-induced catalytic activation. We analyzed the functional importance of the covalent linkage using an isosteric chromophore-protein variant in which the cysteine is replaced by a glycine residue and the chromophore by thiomethyl-p-coumaric acid (TMpCA). The model compound TMpCA is shown to weakly complex with the C69G protein. This non-covalent binding results in considerable tuning of both the pKa and the color of the chromophore. The photoactivity of this system, however, was strongly impaired, making PYP the first known photosensor protein in which the covalent linkage of the chromophore is of paramount importance for the functional activity of the protein in vitro. We also studied the influence of chromophore analogs on the color and photocycle of PYP, not only in WT, but especially in the E46Q mutant, to test if effects from both chromophore and protein modifications are additive. When the E46Q protein binds the sinapinic acid chromophore, the color of the protein is effectively changed from yellow to orange. The altered charge distribution in this protein also results in a changed pKa value for chromophore protonation, and a strongly impaired photocycle. Both findings extend our knowledge of the photochemistry of PYP for signal generation. © The Royal Society of Chemistry and Owner Societies.
Subject
Biology
Biotechnology
4 hydroxy cinnamic acid
apoprotein
cinnamic acid derivative
coumaric acid
cysteine
glycine
mutant protein
photoactive yellow protein
protein
sinapic acid
thiomethyl 4 coumaric acid
article
catalysis
chemical binding
chromatophore
color
covalent bond
in vitro study
knowledge
model
photochemistry
pKa
point mutation
priority journal
protein binding
protein function
protein modification
protein variant
proton transport
Bacterial Proteins
Coumaric Acids
Hydrogen-Ion Concentration
Photoreceptors, Microbial
Point Mutation
Protein Binding
Spectrophotometry, Ultraviolet
Bacteria (microorganisms)
To reference this document use:
http://resolver.tudelft.nl/uuid:427c3f4a-a056-41dc-a698-83077ce29e32
DOI
https://doi.org/10.1039/b701072a
TNO identifier
239979
ISSN
1474-905X
Source
Photochemical and Photobiological Sciences, 6 (5), 571-579
Document type
article