| Budget Amount *help |
¥3,890,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥390,000)
Fiscal Year 2007: ¥1,690,000 (Direct Cost: ¥1,300,000、Indirect Cost: ¥390,000)
Fiscal Year 2006: ¥2,200,000 (Direct Cost: ¥2,200,000)
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| Research Abstract |
Iba1 (ionized calcium-binding adaptor molecule 1) with 147 amino acid residues has been identified as a calcium (Ca^<2+>)-binding protein, expressed specifically in microglia/macrophages, and is expected to be a key factor in membrane ruffling which is a typical feature of activated microglia. We have determined the crystal structure of human Iba1 in Ca^<2+> free form and mouse Iba1 in Ca^<2+> -bound form, to a resolution of 1.9 A and 2.1 A, respectively. X-ray structures of Iba1 revealed a compact, single-domain protein with two EF-hand motifs, showing similarity in overall topology to partial structures of the classical EF-hand proteins troponin C and calmodulin. In mouse Iba1, the second EF-hand contains a bound Ca^<2+>, but the first EF-hand does not, which is often the case in S100 proteins, suggesting that Iba1 has S100 protein-like EF-hands. However, the molecular conformational change induced by Ca^<2+>-binding of Iba1 is different from that found in the classical EF-hand proteins and/or S100 proteins, leading to the formation of a dimer in a crystal, which demonstrates that Iba1 has an unique molecular switching mechanism dependent on Ca^<2+>-binding. The dimer has the shape of a slightly flat quadratic prism with the characteristic charge distribution; negatively charged regions are widely distributed on the upper surface and sides, while the unusual positively charged region is found at the center of the lower surface. It is likely that this positively charged region interacts with a target molecule. To elucidate the oligomeric state of Iba1 in a solution, cross-linking experiments was done, showing that Iba1 partially forms a dimer in a solution. To investigate whether a dimer Iba1 can interacts with target molecules or not, we also done surface plasmon resonance studies, but a clear evidence for this has not yet been obtained.
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