|
ИСТИНА |
Войти в систему Регистрация |
ИСТИНА ПсковГУ |
||
An impact of inter-monomer and inter-domain interactions on the properties of near-infrared fluorescent proteins engineered from bacterial phytochromesтезисы доклада
- Авторы: Stepanenko Olesya V., Stepanenko Olga V., Kuznetsova I.M., Verkhusha V.V., Turoverov K.K.
- Сборник: The FEBS Journal
- Серия: Supplement 1
- Том: 284
- Тезисы
- Год издания: 2017
- Издательство: WILEY-BLACKWELL PUBLISHING
- Местоположение издательства: INC, COMMERCE PLACE, 350 MAIN ST,MALDEN, USA, MA, 02148
- Первая страница: 214
- Последняя страница: 215
- DOI: 10.1111/febs.14174
- Аннотация: Near-infrared fluorescent proteins (NIR FPs) engineered from bacterial phytochromes (BphPs) with the absorbance and fluorescence in the "NIR transparency window" of biological tissues meet the requirements for probes for deep-tissue in vivo imaging. Spectral properties of NIR FPs, composed of two PAS and GAF domains are affected by a position of the covalent attachment of their natural biliverdin chromophore (BV), with the BV-CysPAS adduct being more red-shifted relative to the BV-CysGAF adduct. The covalent binding of BV and its linkage position influences the stability of NIR FP holoprotein. Earlier, covalent binding of the chromophore in dimeric NIR FPs was proposed to depend on inter-monomer and inter-domain interactions. The fraction of the non-covalently bound chromophore in dimeric NIR FPs resulted in decreased quantum yield and brightness of the proteins. Here, we studied the biochemical, structural and spectral properties of a monomeric NIR FP, termed BphP1-FP, and its mutants in buffer solutions and in the presence of denaturant. BphP1-FP has two reactive Cys residues (CysPAS and CysGAF) while its mutants bears either CysPAS or CysGAF, or none of them. All BphP1-FP variants were monomeric. The elimination of inter-monomer interactions led to the absence of the non-covalently bound BV in the BphP1-FP variants. It also confirmed the inter-monomeric allosteric influence on the BV interaction with the monomers in dimeric NIR FPs. The unfolding of monomeric BphP1-FP variants in apoform was irreversible that was in contrast to dimeric NIR FPs, thus indicating that the inter-molecular contacts contribute to folding of NIR FPs. The increased stability of BphP1-FP is likely connected with the inter-domain cross-linking of the PAS and GAF domains to BV. We suggest that similar structural changes in dimeric NIR FPs remove inhibition of the BV covalent binding and increase their stability. This work was supported by the MCB Program of RAS and RFBP grant (16-04-01515).
- Добавил в систему: Степаненко Ольга Викторовна