89745 20210902121805.0 doi 10.3390/ijms21103738 sideral 118161 ART-2020-118161 eng Serrano, A. Universidad de Zaragoza Insights into the fmnat active site of fad synthase: Aromaticity is essential for flavin binding and catalysis 2020 Access copy available to the general public Unrestricted The last step in the biosynthesis of flavin adenine dinucleotide (FAD) is considered a target for the design of antimicrobial drugs because it is carried out by two non-homologous proteins in eukaryotic and prokaryotic organisms. Monofunctional FMN: adenylyltransferases (FMNAT) in Eukarya and FMNAT modules of bifunctional FAD synthases (FADS) in Prokarya belong to different structural families with dissimilar chemistry and binding modes for the substrates. In this study, we analyzed the relevance of the hydrophobic environment of the flavin isoalloxazine in the FMNAT active site of Corynebacterium ammoniagenes FADS (CaFADS) through the mutational analysis of its F62, Y106, and F128 residues. They form the isoalloxazine binding cavity and are highly conserved in the prokaryotic FADS family. The spectroscopic, steady-state kinetics and thermodynamic data presented indicate that distortion of aromaticity at the FMNAT isoalloxazine binding cavity prevents FMN and FAD from correct accommodation in their binding cavity and, as a consequence, decreases the efficiency of the FMNAT activity. Therefore, the side-chains of F62, Y106 and F128 are relevant in the formation of the catalytic competent complex during FMNAT catalysis in CaFADS. The introduced mutations also modulate the activity occurring at the riboflavin kinase (RFK) module of CaFADS, further evidencing the formation of quaternary assemblies during catalysis. info:eu-repo/grantAgreement/ES/DGA-FEDER/E35-20R info:eu-repo/grantAgreement/ES/MINECO-AEI-FEDER/BIO2016-75183-P info:eu-repo/semantics/openAccess by http://creativecommons.org/licenses/by/3.0/es/ 5.923 2020 BIOCHEMISTRY & MOLECULAR BIOLOGY 67 / 297 = 0.226 2020 Q1 T1 CHEMISTRY, MULTIDISCIPLINARY 49 / 178 = 0.275 2020 Q2 T1 1.455 2020 Catalysis 2020 Q1 Computer Science Applications 2020 Q1 Inorganic Chemistry 2020 Q1 Spectroscopy 2020 Q1 Molecular Biology 2020 Q1 Organic Chemistry 2020 Q1 Physical and Theoretical Chemistry 2020 Q1 Medicine (miscellaneous) 2020 Q1 info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion Arilla-Luna, S. Medina, M. Universidad de Zaragoza (orcid)0000-0001-8743-0182 1002 060 Universidad de Zaragoza Dpto. Bioq.Biolog.Mol. Celular Área Bioquímica y Biolog.Mole. 21, 10 (2020), 3738 [16 pp] Int. j. mol. sci. International Journal of Molecular Sciences 1661-6596 1011542 http://zaguan.unizar.es/record/89745/files/texto_completo.pdf Versión publicada 514768 http://zaguan.unizar.es/record/89745/files/texto_completo.jpg?subformat=icon icon Versión publicada oai:zaguan.unizar.es:89745 articulos driver 2021-09-02-09:57:49 ARTICLE