Selective protein photocleavage by fluorescein derivatives.

Modification of the structure of small molecular probe which can act as photocleavage reagent has become a considerable challenge to improve the ability to target specific sites on a large protein. These photoreagents can provide valuable information on the binding site recognition and the mechanism of the photocleavage reaction under photochemical control. In this study, site specific photocleavage of lysozyme and avidin by fluorescein derivatives, fluorescein sodium salt (F-1) and 5(6)-carboxyfluorescein diacetate (F-2) were reported here for the first time. Functional groups on the photoreagent have been proven to effect on the interaction with the protein. Cleavage of the proteins by fluorescein derivatives were successful under visible region when irradiating the solution mixture of protein, fluorescein derivative and electron acceptor, cobalt (III) hexamine trichloride, at 490-492 nm. N-terminal amino acid sequencing of the cleaved fragments of lysozyme indicated the cleavage site between Trp108 - Val 109 for both probes, whereas the cleavage of avidin by F-1 and F-2 were detected between Trp70 - Lys71. Binding interaction can be investigated using methods as simple as absorption and fluorescence spectroscopies. Absorption and fluorescence studies indicated the strong binding interactions between fluorescein derivatives and the target proteins. Computational modeling was used to gain a better insight of the protein-probe binding interaction and binding sites. Molecular docking studies indicated that F-1 and F-2 were located near the hydrophilic and hydrophobic sites of both proteins within 4 Å away from the cleavage site. The docking results clarified the binding sites of F-1 and F-2 on proteins, corresponding to the results obtained from the protein photocleavage studies. Copyright © 2020 Elsevier B.V. All rights reserved.

Citation

Benchawan Jityuti, Mayuso Kuno, Teerayuth Liwporncharoenvong, Apinya Buranaprapuk. Selective protein photocleavage by fluorescein derivatives. Journal of photochemistry and photobiology. B, Biology. 2020 Nov;212:112027

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PMID: 32977112

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