| US 7,579,165 B2 | ||
| Methods for derivatizing protein or peptide with sulfonic acid groups | ||
| Shigemi Norioka, Ibaraki (Japan); Minoru Yamaguchi, Kyoto (Japan); Hiroki Kuyama, Kyoto (Japan); Takashi Obama, Kyoto (Japan); Eiji Ando, Kyoto (Japan); Takashi Nakazawa, Nara (Japan); Norikazu Ueyama, Osaka (Japan); and Taka-aki Okamura, Osaka (Japan) | ||
| Assigned to Shimadzu Corporation, Kyoto (Japan) | ||
| Filed on Oct. 13, 2004, as Appl. No. 10/962,575. | ||
| Claims priority of application No. 2003-356809 (JP), filed on Oct. 16, 2003. | ||
| Prior Publication US 2005/0084927 A1, Apr. 21, 2005 | ||
| Int. Cl. C07K 1/13 (2006.01); C07K 1/14 (2006.01); C07K 14/00 (2006.01); C12P 21/06 (2006.01); C12Q 1/00 (2006.01); C12Q 1/34 (2006.01); C12Q 1/37 (2006.01); G01N 33/53 (2006.01); G01N 33/38 (2006.01); G01N 33/483 (2006.01) | ||
| U.S. Cl. 435—68.1 [435/7.5; 435/18; 435/23; 435/24; 530/300; 530/324; 530/325; 530/326; 530/327; 530/328; 530/329; 530/406] | 17 Claims |
| 1. A method for derivatizing a protein or peptide to a sulfonic acid derivative, comprising steps of:
modifying a N-terminus in a protein or peptide with an immobilized compound A bound to a solid support to obtain a protein
or peptide modified with the immobilized compound A at the N-terminus, said immobilized compound A comprising a disulfide
group and a functional group which reacts with the N-terminus in the protein or peptide;
fragmenting the obtained protein or peptide modified with the immobilized compound A into a N-terminal peptide fragment comprising
the immobilized compound A and containing a disulfide bond, and one or more peptide fragments other than said N-terminal peptide
fragment;
separating the N-terminal peptide fragment from the other peptide fragments to selectively collect the N-terminal peptide
fragment; and
cleaving the disulfide bond of the disulfide group to convert into a sulfonic acid group, thereby converting the modified
protein or peptide into a sulfonic acid derivative,
wherein said immobilized compound A is selected from the group consisting of 3-([2-aminoethyl]dithio)propionic acid hydrochloride
(AEDP), cystine, AEDP in which the carboxyl group of AEDP is replaced by an isothiocyanate or active ester, and cystine in
which the carboxyl group of cystine is replaced by an isothiocyanate or active ester, and
wherein in the cleaving step, the disulfide bond in the N-terminal peptide fragment that originates from the immobilized compound
A is cleaved and converted into the sulfonic acid group and thereby the derivatized N-terminal peptide fragment is obtained.
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