doi:10.1111/ppl.12621engKoskela, M.M.Dahlström, K.M.Goñi, G.Lehtimäki, N.Nurmi, M.Velazquez-Campoy, A.Hanke, G.Bölter, B.Salminen, T.A.Medina, M.Mulo, P.Arabidopsis FNRL protein is an NADPH-dependent chloroplast oxidoreductase resembling bacterial ferredoxin-NADP+ reductasesART-2018-104602Plastidic ferredoxin-NADP+ oxidoreductases (FNRs; EC:1.18.1.2) together with bacterial type FNRs (FPRs) form the plant-type FNR family. Members of this group contain a two-domain scaffold that forms the basis of an extended superfamily of flavin adenine dinucleotide (FAD) dependent oxidoreductases. In this study, we show that the Arabidopsis thaliana At1g15140 [Ferredoxin-NADP+ oxidoreductase-like (FNRL)] is an FAD-containing NADPH dependent oxidoreductase present in the chloroplast stroma. Determination of the kinetic parameters using the DCPIP NADPH-dependent diaphorase assay revealed that the reaction catalysed by a recombinant FNRL protein followed a saturation Michaelis–Menten profile on the NADPH concentration with kcat = 3.2 ± 0.2 s-1, Km NADPH = 1.6 ± 0.3 µM and kcat/Km NADPH = 2.0 ± 0.4 µM-1 s-1. Biochemical assays suggested that FNRL is not likely to interact with Arabidopsis ferredoxin 1, which is supported by the sequence analysis implying that the known Fd-binding residues in plastidic FNRs differ from those of FNRL. In addition, based on structural modelling FNRL has an FAD-binding N-terminal domain built from a six-stranded ß-sheet and one a-helix, and a C-terminal NADP+-binding a/ß domain with a five-stranded ß-sheet with a pair of a-helices on each side. The FAD-binding site is highly hydrophobic and predicted to bind FAD in a bent conformation typically seen in bacterial FPRs.2018http://zaguan.unizar.es/record/16798110.1111/ppl.12621http://zaguan.unizar.es/record/167981oai:zaguan.unizar.es:167981info:eu-repo/grantAgreement/ES/DGA/B18info:eu-repo/grantAgreement/ES/MINECO/BIO2016-75183-PPHYSIOLOGIA PLANTARUM 162, 2 (2018), 177-190All rights reservedhttp://www.europeana.eu/rights/rr-f/info:eu-repo/semantics/openAccess