2006 Fiscal Year Final Research Report Summary
Nobel strategy of structural biology for investigation of membrane proteins-ligands
| Project/Area Number |
14104017
|
|---|
| Research Category |
Grant-in-Aid for Scientific Research (S)
|
| Allocation Type | Single-year Grants |
|---|
| Research Field |
Physical pharmacy
|
|---|
| Research Institution | The University of Tokyo |
|---|
Principal Investigator |
SHIMADA Ichio The University of Tokyo, Graduate School of Pharmaceutical Sciences, Professor, 大学院薬学研究科, 教授 (70196476)
|
|---|
| Co-Investigator(Kenkyū-buntansha) |
TERASAWA Hiroak The University of Tokyo, Graduate School of Medical and Pharmaceutical Sciences Kumamoto University, Professor, 大学院医学薬学研究部, 教授 (10300956)
|
|---|
| Project Period (FY) |
2002 – 2006
|
| Keywords | Stable-isotope labeling / cross-saturation / transferred cross saturation / membrane Protein / interaction / NMR |
|---|
| 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.
|
Research Products
(63 results)
