Probing the spectral signatures of orange carotenoid protein by orthogonal translation with aromatic non-canonical amino acids

• Published in: Biochemical and biophysical research communications 2022-06, Vol.607, p.96-102
• Main Authors: Tseng, Hsueh-Wei, Moldenhauer, Marcus, Friedrich, Thomas, Maksimov, Eugene G., Budisa, Nediljko
• Format: Article
• Language: English
• Subjects: Amino Acids - metabolism; Amino Acids, Aromatic - metabolism; aromatic ncAAs; Bacterial Proteins - metabolism; Carotenoids - metabolism; Cyanobacteria - metabolism; Hydrogen bonding; Methanosarcina mazei pyrrolysyl-tRNA synthetase (MmPylRS); Non-canonical amino acids (ncAAs); Orange carotenoid protein (OCP); site-Specific incorporation; Xanthophyll photoconversion
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1016/j.bbrc.2022.03.118

Orange Carotenoid Protein (OCP) is a water-soluble photoreceptor involved in photoprotection of cyanobacteria. The photoactive OCP contains a bound ketocarotenoid cofactor held in a protein matrix with a hydrogen bonding network. We have developed a system to replace essential residues of the photoactive OCP with non-canonical aromatic analogues that produce well-defined chemical or steric changes. Preliminary spectroscopic evaluation of the generated OCP variants demonstrates the potential of this "molecular surgery" to disentangle protein-chromophore interaction networks that are critical for photoreceptor function. In this way, the number and strength of key contacts with non-canonical amino acids could be controlled and manipulated. We have illustrated this principle here by replacing hydrogen bond donating residues with aromatic non-canonical amino acids that alter the state preference of OCP. •Orange Carotenoid Protein (OCP) is a water-soluble photoreceptor involved in photoprotection of cyanobacteria.•OCP contains a bound ketocarotenoid cofactor held in a protein matrix with a hydrogen bond network.•OCP expression with expanded genetic code enabled the study interactions between the ketocarotenoid and the protein matrix.•The number and strength of key contacts could be controlled and manipulated with non-canonical amino acids.•Replacement of hydrogen-bond donating residues by aromatic non-canonical amino acids changes state preference of OCP..