| Budget Amount *help |
¥3,150,000 (Direct Cost: ¥3,000,000、Indirect Cost: ¥150,000)
Fiscal Year 2007: ¥650,000 (Direct Cost: ¥500,000、Indirect Cost: ¥150,000)
Fiscal Year 2006: ¥2,500,000 (Direct Cost: ¥2,500,000)
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| Research Abstract |
In the present study, we performed the experiment of hydrogen assisted sonolytic hydrolysis of the peptide bond, which resulted in the sequence reflected peptide products with proline information and without any side reactions. The application of ultrasound to an aqueous solution of analyte peptide (ACTH18-39) resulted in the oxidation products M+nO (n=1-6) to an intact peptide molecule M. The peaks corresponding to the products caused by hydrogen radical and acid hydrolysis were observed, and the products were characterized by the C-terminal structures of imidate and carboxylic acid with the mass difference of 1 Da. The result obtained indicates that a hydrolytic sonolysis at peptide bond C-N and hydrogen-assisted cleavage at N-Cα bond occurs competitively by ultrasound in aqueous solutions. In order to enhance the yield of sonolytic products with hydrogen radical, phenolic compounds as hydrogen donor were added to the aqueous solution of the analyte peptide. The addition of 1,2-benzenediol to the aqueous solution of ACTH18-39 brought about a considerable enhancement of the yield of product originated from the sonolytic hydrolysis at peptide bonds. It was clear that the addition of 1,2-benzenediol as a hydrogen donor did not enhance the hydrogen radical-initiated cleavage at N-Cα bond, but the sonolytic hydrolysis of peptide bonds. The mass difference between adjacent peaks was equal to the mass of amino acid residues, -NH-CαH (R) -CO-, on the peptide backbone, so that sequence could be easily determined from the spectrum. Any interfering peaks originated from side reactions such as dehydration, oxidation and reduction were little observed.
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