Partially Folded Structure of Flavin Adenine Dinucleotide-depleted Ferredoxin-NADP+ Reductase with Residual NADP+ Binding Domain

This research was originally published in the Journal of Biological Chemistry. Masahiro Maeda, Daizo Hamada, Masaru Hoshino, Yayoi Onda, Toshiharu Hase and Yuji Goto. Partially Folded Structure of Flavin Adenine Dinucleotide-depleted Ferredoxin-NADP+ Reductase with Residual NADP+ Binding Domain. J. Biol. Chem. 2002; 277, 17101-17107. © the American Society for Biochemistry and Molecular Biolog

Cores and pH-dependent Dynamics of Ferredoxin-NADP+ Reductase Revealed by Hydrogen/Deuterium Exchange

This research was originally published in the Journal of Biological Chemistry. Young-Ho Lee, Kosuke Tamura, Masahiro Maeda, Masaru Hoshino, Kazumasa Sakurai, Satoshi Takahashi, Takahisa Ikegami, Toshiharu Hase, and Yuji Goto. Cores and pH-dependent Dynamics of Ferredoxin-NADP+ Reductase Revealed by Hydrogen/Deuterium Exchange. J. Biol. Chem. 2007; 282, 5959-5967. © the American Society for Biochemistry and Molecular Biolog

Intracellular Trafficking of the Amyloid β-Protein Precursor (APP) Regulated by Novel Function of X11-Like

Background: Amyloid beta (A beta), a causative peptide of Alzheimer's disease, is generated by intracellular metabolism of amyloid beta-protein precursor (APP). In general, mature APP (mAPP, N- and O-glycosylated form) is subject to successive cleavages by alpha- or beta-, and gamma-secretases in the late protein secretory pathway and/or at plasma membrane, while immature APP (imAPP, N-glycosylated form) locates in the early secretory pathway such as endoplasmic reticulum or cis-Golgi, in which imAPP is not subject to metabolic cleavages. X11-like (X11L) is a neural adaptor protein composed of a phosphotyrosine-binding (PTB) and two C-terminal PDZ domains. X11L suppresses amyloidogenic cleavage of mAPP by direct binding of X11L through its PTB domain, thereby generation of A beta lowers. X11L expresses another function in the regulation of intracellular APP trafficking. Methodology: In order to analyze novel function of X11L in intracellular trafficking of APP, we performed a functional dissection of X11L. Using cells expressing various domain-deleted X11L mutants, intracellular APP trafficking was examined along with analysis of APP metabolism including maturation (O-glycosylation), processing and localization of APP. Conclusions: X11L accumulates imAPP into the early secretory pathway by mediation of its C-terminal PDZ domains, without being bound to imAPP directly. With this novel function, X11L suppresses overall APP metabolism and results in further suppression of Ab generation. Interestingly some of the accumulated imAPP in the early secretory pathway are likely to appear on plasma membrane by unidentified mechanism. Trafficking of imAPP to plasma membrane is observed in other X11 family proteins, X11 and X11L2, but not in other APP-binding partners such as FE65 and JIP1. It is herein clear that respective functional domains of X11L regulate APP metabolism at multiple steps in intracellular protein secretory pathways

Non-covalent forces tune the electron transfer complex between ferredoxin and sulfite reductase to optimize enzymatic activity.

Although electrostatic interactions between negatively-charged ferredoxin (Fd) and positively-charged sulfite reductase (SiR) have been predominantly highlighted to characterize complex formation, the detailed nature of intermolecular forces remains to be fully elucidated. We herein investigated interprotein forces for formation of an electron-transfer complex between Fd and SiR and their relationship to SiR activity using various approaches over NaCl concentrations between 0 and 400 mM. Fd-dependent SiR activity assays revealed a bell-shaped activity curve with a maximum around 40-70 mM NaCl and a reverse bell-shaped dependence of interprotein affinity. Meanwhile, intrinsic SiR activity, as measured in a methyl viologen-dependent assay, exhibited saturation above 100 mM NaCl. Thus, two assays suggested that interprotein interaction is crucial in controlling Fd-dependent SiR activity. Calorimetric analyses showed the monotonic decrease in interprotein affinity on increasing NaCl concentrations, distinguished from a reverse bell-shaped interprotein affinity observed from Fd-dependent SiR activity assay . Furthermore, Fd:SiR complex formation and interprotein affinity were thermodynamically adjusted by both enthalpy and entropy through electrostatic and non-electrostatic interactions. A residue-based NMR investigation on addition of SiR to 15N-labeled Fd at the various NaCl concentration also demonstrated that a combination of electro- and non-electrostatic forces stabilized the complex with similar interfaces and modulated the binding affinity and mode. Our findings elucidate that non-electrostatic forces are also essential for the formation and modulation of the Fd:SiR complex. We suggest that a complex configuration optimized for maximum enzymatic activity near physiological salt conditions is achieved by structural rearrangement through controlled non-covalent interprotein interactions