Nanomechanical Study of Enzyme: Coenzyme Complexes: Bipartite Sites in Plastidic Ferredoxin-NADP+ Reductase for the Interaction with NADP+

Pérez-Domínguez, S. ; Caballero-Mancebo, S. ; Marcuello, C. ; Martínez-Júlvez, M. (Universidad de Zaragoza) ; Medina, M. (Universidad de Zaragoza) ; Gracia Lostao, A.

Resumen: Plastidic ferredoxin-NADP+ reductase (FNR) transfers two electrons from two ferredoxin or flavodoxin molecules to NADP+, generating NADPH. The forces holding the Anabaena FNR:NADP+ complex were analyzed by dynamic force spectroscopy, using WT FNR and three C-terminal Y303 variants, Y303S, Y303F, and Y303W. FNR was covalently immobilized on mica and NADP+ attached to AFM tips. Force–distance curves were collected for different loading rates and specific unbinding forces were analyzed under the Bell–Evans model to obtain the mechanostability parameters associated with the dissociation processes. The WT FNR:NADP+ complex presented a higher mechanical stability than that reported for the complexes with protein partners, corroborating the stronger affinity of FNR for NADP+. The Y303 mutation induced changes in the FNR:NADP+ interaction mechanical stability. NADP+ dissociated from WT and Y303W in a single event related to the release of the adenine moiety of the coenzyme. However, two events described the Y303S:NADP+ dissociation that was also a more durable complex due to the strong binding of the nicotinamide moiety of NADP+ to the catalytic site. Finally, Y303F shows intermediate behavior. Therefore, Y303, reported as crucial for achieving catalytically competent active site geometry, also regulates the concerted dissociation of the bipartite nucleotide moieties of the coenzyme. © 2022 by the authors. Licensee MDPI, Basel, Switzerland.

Idioma: Inglés

DOI: 10.3390/antiox11030537

Año: 2022

Publicado en: Antioxidants 11, 3 (2022), 537 [20 pp]
ISSN: 2076-3921

Factor impacto JCR: 7.0 (2022)

Categ. JCR: BIOCHEMISTRY & MOLECULAR BIOLOGY rank: 46 / 285 = 0.161 (2022) - Q1 - T1
Categ. JCR: FOOD SCIENCE & TECHNOLOGY rank: 13 / 142 = 0.092 (2022) - Q1 - T1
Categ. JCR: CHEMISTRY, MEDICINAL rank: 6 / 60 = 0.1 (2022) - Q1 - T1

Factor impacto CITESCORE: 8.8 - Biochemistry, Genetics and Molecular Biology (Q1)

Factor impacto SCIMAGO: 1.084 - Biochemistry (Q1) - Clinical Biochemistry (Q1) - Food Science (Q1) - Physiology (Q1) - Molecular Biology (Q2) - Cell Biology (Q2)

Financiación: info:eu-repo/grantAgreement/ES/DGA-FEDER/E35-20R
Financiación: info:eu-repo/grantAgreement/ES/MICINN-AEI/PID2019-103901GB-I00

Tipo y forma: Article (Published version)

Área (Departamento): Área Bioquímica y Biolog.Mole. (Dpto. Bioq.Biolog.Mol. Celular)