Biology subjects

Senda, M.

Publications and source records attributed to Senda, M..

3 recordsLinked to original sources

bioRxiv2026

The 329HHK331 motif is essential for Alzheimer's disease tau filament fold

Cryo-electron microscopy (cryo-EM) has revealed disease-specific tau filament folds, yet the local sequence elements that determine them remain poorly understood. Here we focused on the 329HHK331 motif near an inter-protofilament interface in Alzheimers disease (AD)-type tau filaments, and analyzed recombinant dGAE filaments of wild-type (WT) and mutants in this motif. All mutants formed amyloid-like filaments in vitro, but their morphologies differed. In cultured cells, WT filaments efficiently seeded WT tau aggregation. Filaments with three-residue changes (deletion or Ala substitution) showed almost no seeding activity, whereas two-residue deletions retained partial activity. Cryo-EM showed that WT dGAE filaments form a quadruple helical filament of two protofilament dimers. Each dimer comprises two C-shaped protofilaments, centered on a 333GGG335-mediated inter-protofilament interaction and supported by flanking 329HHK331-336QVE338 contacts. Three-residue alterations abolished interactions with the QVE motif at the protofilament interface, thereby displacing 333GGG335 and forming non-C-shaped protofilament structures that are intrinsically poor templates for tau seeding. By contrast, two-residue deletions maintained the C-shaped protofilament structure because the remaining residue formed alternative inter-protofilament interactions. These findings suggest that the 329HHK331 region is a key determinant of the AD-like C-shaped protofilament fold and link this motif to tau seeding, providing insight into disease-specific tau filament formation.

neuroscience↗

bioRxiv2024

Structural characterization of pyruvic oxime dioxygenase, a key enzyme of heterotrophic nitrification

Nitrification by heterotrophic microorganisms is an important part of the nitrogen cycle in the environment. The enzyme responsible for the core function of heterotrophic nitrification is pyruvic oxime dioxygenase (POD). POD is a non-heme Fe(II)-dependent enzyme that catalyzes the dioxygenation of pyruvic oxime to produce pyruvate and nitrite. To analyze the catalytic mechanism of POD, the crystal structure of POD from Alcaligenes faecalis (AfPOD) was determined at 1.76 [A] resolution. The enzyme is a homo-tetramer and the subunit structure is homologous to those of class II aldolases, in particular a zinc-dependent L-fuculose-1-phosphate aldolase. The active site of the subunit is located at the bottom of a cleft formed with an adjacent subunit. The iron ion at the active site is coordinated by three histidines and three water molecules in an octahedral geometry. The putative oxygen tunnel was connected between the active site and the central cavity of the tetramer. The N-terminal region of AfPOD, which is essential for catalytic activity, is disordered in the crystal. Structure prediction with AlphaFold2 combined with mutational experiments suggested that the disordered N-terminal region adopts an -helix conformation and participates in the formation of the active site. The catalytic mechanism of the dioxygenase reaction by POD is discussed on the basis of the molecular docking model. IMPORTANCEOur knowledge of nitrification has increased considerably in recent decades with the discovery of new nitrifying microorganisms and the characterization of their biochemical processes. Some heterotrophic bacteria and fungi are known to show nitrification activities, but the molecular mechanisms had been poorly understood. Here, we performed a structural characterization of POD, a key enzyme in heterotrophic nitrification that produces nitrite from ammonia using pyruvic oxime as an intermediate. Structural and enzymatic analyses revealed that POD is a unique dioxygenase with features such as an aldolase backbone, an N-terminal -helix, and an oxygen tunnel. Our results provide insights not only into the molecular mechanisms but also into the design of specific inhibitors of heterotrophic nitrification.

microbiology↗

bioRxiv2020

Activity-specificity trade-off gives PI5P4Kβ a nucleotide preference to function as a GTP-sensing kinase.

Most kinases function with ATP. However, contrary to the prevailing dogma, phosphatidylinositol 5-phosphate 4-kinase {beta} (PI5P4K{beta}) utilizes GTP as a primary phosphate donor with a unique binding mode for GTP. Although PI5P4K{beta} is evolved from a primordial ATP-utilizing enzyme, PI4P5K, how PI5P4K{beta} evolutionarily acquired the GTP preference to function as a cellular GTP sensor remains unclear. In this study, we show that the short nucleotide base-recognition motif, TRNVF, is responsible for the GTP binding of PI5P4K{beta}, and also confers onto PI5P4K{beta} an unexpected specificity that extends to inosine triphosphate (ITP) and xanthosine triphosphate (XTP). A mutational study with GTP analogues suggests that the extended specificity is an obligatory consequence to the acquisition of GTP-dependent activity. However, as the cellular concentrations of ITP and XTP are typically negligible, PI5P4K{beta} can still function as a GTP sensor, suggesting that the cellular physiological conditions leave room for the functional evolution of PI5P4K{beta}.

biochemistry↗