Total enzymatic synthesis of cholecystokinin CCK-5.
This paper describes the enzymatic synthesis of the C-terminal fragment H-Gly-Trp-Met-Asp-Phe-NH2 of cholecystokinin. Immobilized enzymes were used for the formation of all peptide bonds except thermolysin. Beginning the synthesis with phenylacetyl (PhAc) glycine carboxamidomethyl ester (OCam) and H-Trp-OMe by using immobilized papain as biocatalyst in buffered ethyl acetate, the dipeptide methyl ester was then coupled directly with Met-OEt.HCl by alpha-chymotrypsin/Celite 545 in a solvent free system. For the 3+2 coupling PhAc-Gly-Trp-Met-OEt had to be converted into its OCam ester. The other fragment H-Asp(OMe)-Phe-NH2 resulted from the coupling of Cbo-Asp(OMe)-OH with H-Phe-NH2.HCl and thermolysin as catalyst, followed by catalytic hydrogenation. Finally PhAc-Gly-Trp-Met-Asp-Phe-NH2 was obtained in a smooth reaction from PhAc-Gly-Trp-Met-OCam and H-Asp(OMe)-Phe-NH2 with alpha-chymotrypsin/Celite 545 in acetonitrile, followed by basic hydrolysis of the beta-methyl ester. The PhAc-group is removed with penicillin G amidase and CCK-5 is obtained in an overall isolated yield of 19.6%.
Xiang
H
H
School of Pharmacy, China Pharmaceutical University, Nanjing, P.R. China.
Xiang
G Y
GY
Lu
Z M
ZM
Guo
L
L
Eckstein
H
H
D000085:Acetates / Q000737:chemistry; D001407:Bacillus / Q000378:metabolism; D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D004800:Enzymes, Immobilized / Q000737:chemistry*; D004952:Esters / Q000737:chemistry; D005998:Glycine / Q000737:chemistry; D006868:Hydrolysis; D010206:Papain / Q000737:chemistry; D010399:Penicillin Amidase / Q000737:chemistry; D010400:Penicillin G / Q000737:chemistry; D010418:Pentagastrin / Q000737:chemistry; D010446:Peptide Fragments / Q000138:chemical synthesis* / Q000737:chemistry*; D010455:Peptides / Q000737:chemistry; D012844:Sincalide / Q000138:chemical synthesis* / Q000737:chemistry*; D012997:Solvents / Q000737:chemistry; D013820:Thermolysin / Q000737:chemistry
D000085:Acetates; D004800:Enzymes, Immobilized; D004952:Esters; D010446:Peptide Fragments; D010455:Peptides; D012997:Solvents; C049128:cholecystokinin pentapeptide; C007650:ethyl acetate; D010206:Papain; D013820:Thermolysin; D010399:Penicillin Amidase; D010418:Pentagastrin; D012844:Sincalide; D010400:Penicillin G; D005998:Glycine