| Budget Amount *help |
¥118,560,000 (Direct Cost: ¥91,200,000、Indirect Cost: ¥27,360,000)
Fiscal Year 2006: ¥18,720,000 (Direct Cost: ¥14,400,000、Indirect Cost: ¥4,320,000)
Fiscal Year 2005: ¥18,720,000 (Direct Cost: ¥14,400,000、Indirect Cost: ¥4,320,000)
Fiscal Year 2004: ¥18,720,000 (Direct Cost: ¥14,400,000、Indirect Cost: ¥4,320,000)
Fiscal Year 2003: ¥18,720,000 (Direct Cost: ¥14,400,000、Indirect Cost: ¥4,320,000)
Fiscal Year 2002: ¥43,680,000 (Direct Cost: ¥33,600,000、Indirect Cost: ¥10,080,000)
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| Research Abstract |
1)Development of NMR method for lager proteins :
In the previous paper, it has been shown that the cross-saturation method enables us to identify the contact. residues of large protein complexes in a more rigorous manner than chemical shift perturbation and hydrogen-deuterium exchange experiments [Nat.Struct.Biol.7(3)220-223]. However, within the determination of the contact residues by the cross-saturation method, there are limitations that the method is difficult to apply to protein complexes with a molecular weight. over 150 K and/or with weak binding, since the resonances originating from the complexes should be directly observed in the method. In the present research. to overcome these limitations, we carried out the cross-saturation measurements under the condition of a fast exchange between free and bound states on the NMR time scale, and determined the contact residues of the complex of the B domain of protein A and intact. IgG which has a molecular weight of 164 K and shows weak binding.
2)Interaction analyses between an ion channel and its pore blocker
We have determined the binding site on agitoxin2 (AgTx2) to the KcsA K+ channel by a transferred cross-saturation (TCS) experiment. The residues significantly affected in the TCS experiments formed a contiguous surface on AgTx2, and substitutions of the surface residues decreased the binding affinity to the KcsA K+ channel. Based on properties of the AgTx2 binding site with the KcsA K+ channel, we present a surface motif that is observed on pore-blocking toxins affecting the K+ channel. Furthermore, we also explain the structural basis of the specificity of the K+ channel to the toxins. The TCS method utilized here is applicable not only for the channels. which are complexed with other inhibitors but also with a variety of regulatory molecules, and provides important information about their interface in solution.
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