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SUNTORY FOUNDATION FOR LIFE SCIENCES




Oct.2007

Research Summary

AVANCE-750 NMR , apex-94Qe FT-ICR MS

Recent great progresses in analytical methods and instrumentations have enabled determination of precise molecular structures and conformations, leading to deeper understandings of their functions in physiological events, and to development of new applications of the molecules.

During these decades we have been accumulating knowledge and techniques for determination of molecular structures of secondary metabolites and natural products with significant biological functions. The structures of those molecules which are obtained in extremely small amounts from natural sources are also studied. In addition to such relatively small natural products, we are now studying their target proteins such as receptors and transporters as well as their interactions with lipid membranes. Dynamic aspects of protein interactions have also become our important research subjects.


Major Research Projects

1) Investigation of the initial events of innate immune recognition played by Toll-like receptor 4, MD-2 and lipopolysaccharide

pathway of innate immune mechanism

Lipopolysaccharides from gram-negative bacteria such as Escherichia coli cause fever and even fatal diseases such as septic shock. Toll-like receptor 4 (TLR4), identified recently as a receptor recognizing lipopolysaccharides, is now attracting attention as an important member of innate immune system that prevents the expansion of early infections. TLR4 recognizes lipopolysaccharides via a cofactor protein known as MD-2, and then forms a dimer, resulting in the transduction of innate immune signals into cells. Despite the attention, much remains unknown about details of the molecular mechanism of the initial innate immune events. We have been working to analyze lipopolysaccharides in phospholipids, and TLR4/MD-2/lipopolysaccharide interactions with a focus on the action mechanism of lipopolysaccharides on living organisms.

Our interest also goes to the mechanism of discrimination between self and non-self in the innate immune system. In this theme, we apply NMR relaxation spectroscopy, a powerful tool we have recently developed, to analyze the dynamics of the molecular interaction to elucidate the mechanism of the innate immune recognition.

  1. Interaction of Lipopolysaccharide and Phospholipid in Mixed Membranes: Solid-State 31P-NMR Spectroscopic and Microscopic Investigations.
    Nomura K., Inaba T., Morigaki K., Brandenburg K., Seydel U., Kusumoto S.
    Biophys J. (2008) in press.
  2. Mechanism of coupled folding and binding of an intrinsically disordered protein.
    Sugase K., Dyson H. J., Wright P. E.
    Nature 447 (7147), 1021-1025 (2007).
  3. Tailoring relaxation dispersion experiments for fast-associating protein complexes.
    Sugase K., Lansing J. C., Dyson H. J., Wright P. E.
    J. Am. Chem. Soc. 129 (44), 13406-13407 (2007).

2) Structural analysis of Fe(III)-mugineic acid (MA) complex and its transporter HvYS1

Crystal Structure of MA-Cu Complex

The Fe(III)-mugineic acid complex plays a key role in the growth of gramineous plants. Even after nearly 30 years since its discovery, the 3-dimensional structure of the complex remains undetermined, despite the fact that the 3-dimensional structures of the corresponding complexes of Co(III) and Cu(II) are already known. This is because no crystals of the iron complex have been available. Hence, we are working to analyze the complex in solution using paramagnetic NMR spectroscopy. Regarding the transporter protein involved in the uptake of iron-MA complex by plants, we investigate the correlation between the ligand selectivity and the 3-dimensional structure of the outer membrane region of the protein.

Meanwhile, we are attempting to analyze the form of iron existing in root cells after absorption and how it is transported using an ultra-high-resolution FT-ICR MS instrument by the aid of a newly elaborated method.

  1. Identification of Two Novel Phytosiderophores Secreted by Perennial Grasses.
    Ueno D., Rombolà A. D., Iwashita T., Nomoto K., Ma J. F.
    New Phytol. 174 (4), 304-310 (2007).

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