![]() FNRhq transfers then both electrons in a single hydride transfer step to NADP(+). The FAD cofactor of FNR accepts two electrons from two independent reduced ferredoxin molecules (Fd) in two sequential steps, first producing neutral semiquinone and then the fully anionic reduced, or hydroquinone, form of the enzyme (FNRhq). 1).įerredoxin-NADP(+) reductase (FNR) catalyzes the last step of linear electron transfer in photosynthetic light reactions. These studies indicated that the mutual disposition between Fd and FNR after the first encounter is not optimal and that the charged residues on the surfaces of Fd and FNR provide a dipole moment that helps their mutual relative orientation and makes their first encounter possible (Fig. 1a-c) are crucial in modulating the midpoint reduction potential and stabilizing the semiquinone, a key factor for efficient transfer of one electron between Fd and FNR (Aliverti et al. ![]() Moreover, FNR active site residues, namely S80, E301, and the C-terminal Y303, as well as some residues at the FADbinding domain, namely R16, L76, L78, and particularly K75 (Anabaena numbering, Fig. Combination of these methodologies identified key residues at the interaction surfaces of these proteins as well as in the modulation of their redox centers, ascribing major and specific functions to some of them as reviewed elsewhere (Hurley et al. Our results probe that the active site of FNR is tuned by a H-bond network that involves the side-chains of these residues and that results critical for optimal substrate binding, exchange of electrons and, particularly, competent disposition of the C4n (hydride acceptor/donor) of the nicotinamide moiety of the coenzyme during the reversible HT event. ![]() Additionally, we revise the role of Tyr79 and Ser80, previously investigated in homologous enzymes from plants. We show that Ser59 indirectly modulates the geometry of the active site, the interaction with substrates and the electronic properties of the isoalloxazine ring, and in consequence the ET and HT processes. Here, we analyse for the first time the role of Ser59 in Anabaena FNR, a residue suggested by recent theoretical simulations as putatively involved in competent binding of the coenzyme in the active site by cooperating with Ser80. Despite the good knowledge of this catalytic machinery, additional roles can still be envisaged for already reported key residues, and new features are added to residues not previously identified as having a particular role in the mechanism. In addition, its charge modulates the two one-electron redox potentials of the flavin to stabilize the semiquinone form.įerredoxin-NADP(+) reductase (FNR) catalyses the production of NADPH in photosynthetic organisms, where its FAD cofactor takes two electrons from two reduced ferredoxin (Fd) molecules in two sequential steps, and transfers them to NADP(+) in a single hydride transfer (HT) step. These functional and structural studies lead us to conclude that Glu-312 does not fulfil the role of proton donor during catalysis, but it is required for proper binding of the nicotinamide ring of NADP(H). Furthermore, NADP(H) binding was partially perturbed. Rapid kinetic absorption spectroscopy studies demonstrated that flavin reduction by NADPH was impaired in the mutants. The E312L mutant was the only one that was almost inactive (approximately 1%), whereas unexpectedly the E312A reductase was 10-100% active with the various acceptors tested. ![]() Four mutants of the enzyme, in which Glu-312 was replaced with Asp, Gln, Leu, and Ala to probe the role of the residue charge, size, and polarity in the enzyme activity, have been heterologously expressed, purified, and characterized through steady-state, rapid kinetic studies, ligand-binding experiments, and three-dimensional structure determination by x-ray crystallography. I suggest the searching by manufacturer to get to the right part of the page.Ferredoxin-NADP+ reductase, the prototype of a large family of structurally related flavoenzymes, pairs single electrons carried by ferredoxin I and transfers them as a hydride to NADP+. It’s all laid out in a very large, 1-page table. This page has a lot of guitar amplifier and distortion pedal schematics on it.
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