title: Energetic basis on interactions between ferredoxin and ferredoxin NADP+reductase at varying physiological conditions.
abstract: In spite of a number of studies to characterize ferredoxin (Fd):ferredoxin NADP+reductase (FNR) interactions at limited conditions, detailed energetic investigation on how these proteins interact under near physiological conditions and its linkage to FNR activity are still lacking. We herein performed systematic Fd:FNR binding thermodynamics using isothermal titration calorimetry (ITC) at distinct pH (6.0 and 8.0), NaCl concentrations (0-200 mM), and temperatures (19-28 °C) for mimicking physiological conditions in chloroplasts. Energetically unfavorable endothermic enthalpy changes were accompanied by Fd:FNR complexation at all conditions. This energetic cost was compensated by favorable entropy changes, balanced by conformational and hydrational entropy. Increases in the NaCl concentration and pH weakened interprotein affinity due to the less contribution of favorable entropy change regardless of energetic gains from enthalpy changes, suggesting that entropy drove complexation and modulated affinity. Effects of temperature on binding thermodynamics were much smaller than those of pH and NaCl. NaCl concentration and pH-dependent enthalpy and heat capacity changes provided clues for distinct binding modes. Moreover, decreases in the enthalpy level in the Hammond's postulate-based energy landscape implicated kinetic advantages for FNR activity. All these energetic interplays were comprehensively demonstrated by the driving force plot with the enthalpy-entropy compensation which may serve as an energetic buffer against outer stresses. We propose that high affinity at pH 6.0 may be beneficial for protection from proteolysis of Fd and FNR in rest states, and moderate affinity at pH 8.0 and proper NaCl concentrations with smaller endothermic enthalpy changes may contribute to increase FNR activity.
authors.affiliation: Institute for Protein Research, Osaka University, Yamadaoka 3-2, Suita, Osaka, 565-0871, Japan.
authors.forename: Misaki
authors.initials: M
authors.lastname: Kinoshita
authors.affiliation: Institute for Protein Research, Osaka University, Yamadaoka 3-2, Suita, Osaka, 565-0871, Japan.
authors.forename: Ju Yaen
authors.initials: JY
authors.lastname: Kim
authors.affiliation: Cellular Function Imaging Team, Division of Bio-function Dynamics Imaging, RIKEN Center for Life Science Technologies, Kobe, Hyogo, 650-0047, Japan.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Kume
authors.affiliation: Institute for Protein Research, Osaka University, Yamadaoka 3-2, Suita, Osaka, 565-0871, Japan.
authors.forename: Yuxi
authors.initials: Y
authors.lastname: Lin
authors.affiliation: Trinity College Dublin, Trinity Biomedical Sciences Institute (TBSI), School of Biochemistry & Immunology, Dublin 2, Ireland.
authors.forename: K Hun
authors.initials: KH
authors.lastname: Mok
authors.affiliation: Cellular Function Imaging Team, Division of Bio-function Dynamics Imaging, RIKEN Center for Life Science Technologies, Kobe, Hyogo, 650-0047, Japan.
authors.forename: Yosky
authors.initials: Y
authors.lastname: Kataoka
authors.affiliation: Department of Chemistry, Faculty of Science, Hokkaido University, Sapporo, Japan.
authors.forename: Koichiro
authors.initials: K
authors.lastname: Ishimori
authors.affiliation: Malvern Instrument Inc., 22 Industrial Drive East, Northampton, MA, 01060, USA.
authors.forename: Natalia
authors.initials: N
authors.lastname: Markova
authors.affiliation: Institute for Protein Research, Osaka University, Yamadaoka 3-2, Suita, Osaka, 565-0871, Japan.
authors.forename: Genji
authors.initials: G
authors.lastname: Kurisu
authors.affiliation: Institute for Protein Research, Osaka University, Yamadaoka 3-2, Suita, Osaka, 565-0871, Japan.
authors.forename: Toshiharu
authors.initials: T
authors.lastname: Hase
authors.affiliation: Institute for Protein Research, Osaka University, Yamadaoka 3-2, Suita, Osaka, 565-0871, Japan. Electronic address: mr0505@protein.osaka-u.ac.jp.
authors.forename: Young-Ho
authors.initials: YH
authors.lastname: Lee
mesh_terms: D019277:Entropy; D005287:Ferredoxin-NADP Reductase / Q000378:metabolism*; D005288:Ferredoxins / Q000378:metabolism*; D007700:Kinetics; D018515:Plant Leaves / Q000378:metabolism*; D010940:Plant Proteins / Q000378:metabolism*; D011485:Protein Binding; D012965:Sodium Chloride / Q000378:metabolism; D013816:Thermodynamics; D003313:Zea mays / Q000378:metabolism*
chemical_list: D005288:Ferredoxins; D010940:Plant Proteins; D012965:Sodium Chloride; D005287:Ferredoxin-NADP Reductase
keywords: Binding thermodynamics; Calorimetry; Driving force plot; Electron transfer complex; Ferredoxin; Ferredoxin-NADP(+) reductase