Synthesis of N alpha-protected aminoacyl 7-amino-4-methyl-coumarin amide by phosphorous oxychloride and preparation of specific fluorogenic substrates for papain.
We report an improved procedure for the synthesis of fully protected aminoacyl 7-amino-4-methylcoumarin amide (MCA) employing the phosphorous oxychloride anhydride method. Seven Boc-X-MCA [where X = Arg(NG Tos), Cys(S-Bzl), Thr(O-Bzl), Ser(O-Bzl), Phe, Leu and Gly] and Z-Tyr(O-Me) were synthesized using this procedure, with yields ranging from 50% to 75%. These aminoacyl-MCA derivatives were employed for the synthesis of epsilon-NH2-caproyl-Leu-X-MCA, a fluorescent peptide series, which were assayed as papain substrates. All of them were completely hydrolyzed by papain, indicating that all of the Boc-X-MCA derivatives obtained were practically free of racemization. Since epsilon-NH2-Caproyl-Leu-(S-Bzl)Cys-MCA is very susceptible to hydrolysis by papain, quite resistant to hydrolysis by chymotrypsin and not hydrolyzed by trypsin, it is recommended for assays of thiol-proteinases in which specificity is required.
Alves
L C
LC
Escola Paulista de Medicina, São Paulo, Brazil.
Almeida
P C
PC
Franzoni
L
L
Juliano
L
L
Juliano
M A
MA
D000596:Amino Acids / Q000737:chemistry*; D003374:Coumarins / Q000737:chemistry*; D004151:Dipeptides / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000138:chemical synthesis*; D010206:Papain / Q000378:metabolism*; D010758:Phosphorus / Q000737:chemistry; D017553:Phosphorus Compounds
D000596:Amino Acids; D003374:Coumarins; D004151:Dipeptides; D005456:Fluorescent Dyes; D017553:Phosphorus Compounds; D010758:Phosphorus; C013196:phosphoryl chloride; D010206:Papain