| Budget Amount *help |
¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 2003: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 2002: ¥1,500,000 (Direct Cost: ¥1,500,000)
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| Research Abstract |
1)X-ray crystal structure analyses of C-terminal amidated and nonamidated dipeptides
As a series of elucidating the structural features of peptides caused by the C-terminal α-amidation, the crystal structures of H-Val-Gly-NH2, H-Ser-Phe-NH2, H-Gly-Tyr-NH2, and H-Pro-Tyr-NH2 hydrochloride salts were analyzed by X-ray diffraction method. Although respective molecules take the energetically allowable torsion angles concerning the backbone and side chains, their conformations are not necessarily the same as the corresponding unamidated ones. This is resulted from the different molecular packing requirement, rather than the different conformational feature inherent in the C-amidated and -unamidated peptides. As for the molecular packing feature, each peptide tends to form the repeated structure through the hydrogen bonds in which both amide NH and O=C groups participate. The chloride ions are located between the neighboring peptides and are hydrogen-bonded to the respective amide NHs, leadin
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g to the sheet structure. The hydrogen-bonding feature of amide group and its function for the molecular packing has been discussed based on the results so far analyzed.
2)Analyses of solution structures of m-opioid agonist endomorphin 2 and its C-terminal OH form
In order to make clear the structural role of the C-terminal amide group of endomorphin-2 (EM2, H-Tyr-Pro-Phe-Phe-NH2), an endogenous m-receptor ligand, for the biological function, the solution conformations of endomorphin-2 and its C-terminal free acid (EM2OH, H-Tyr-Pro-Phe-Phe-OH), studied by two-dimensional ^1H-NMR measurements and molecular modeling calculations, were compared. Both peptides were in equilibrium between the cis and trans isomers around the Tyr-Pro w bond in a population of approximately 1:2 ratio in TFE and water and only trans in the membrane-mimetic micelles of perdeuterated dodecylphosphocholine (DPC). Fifty possible 3D structures for the each isomer were generated by the dynamical simulated annealing method under the proton-proton distance constraints derived from the ROE cross peaks. EM2 conformers adopt an open conformation in both TFE and water, whereas EM2OH shows conformational variation between the extended and folded backbone structures. On the other hand, EM2 in DPC takes the extended and folded backbone structures in 2:1 ratio, whereas EM2OH shows an open conformation. These results indicate clearly that the substitution of carboxyl group for C-terminal amide group makes the peptide flexible. The conformational requirement for μ-receptor activation has been discussed based on the active form proposed for endomorphin-1 and by comparing conformational features of EM2 and EM2OH. Less
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