Discrimination of recombinant and pituitary-derived bovine and porcine growth hormones by peptide mass mapping.
Somatotropins, which are used in cattle for growth and lactating performances, are difficult to reliably detect because no direct method exists. Reversed-phase high-performance liquid chromatography (RP-HLC) coupled to electrospray ionization quadrupole mass spectrometry (ESI/MS) has been developed to separate and characterize the N-terminal peptides resulting from tryptic cleavage of natural and recombinant growth hormones from different species (bovine, porcine, and human) and suppliers. Conditions for tryptic digestion were optimized. This technique was found to be optimal to cleave efficiently the N-terminal peptide of the proteins without releasing too much noise from the matrix. Characterization of the peptides through ESI(+)-MS allowed natural and recombinant growth hormones from bovine and porcine species with N-terminal amino acid sequences differing from one amino acid residue to be discriminated. However, the studied human growth hormones had similar primary sequences that did not permit any discrimination between recombinant and natural forms, thus confirming the known identity of these hormones. Protein digestions with pepsin and chymotrypsin were also compared but were not conclusive due to the too small N-terminal peptides released after proteolysis.
Pinel
Gaud
G
LABERCA, Ecole Nationale Vétérinaire de Nantes, Route de Gachet, B.P. 50707, 44307 Nantes Cedex 3, France. pinel@vet-nantes.fr
André
François
F
Le Bizec
Bruno
B
D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D002918:Chymotrypsin / Q000378:metabolism; D013006:Growth Hormone / Q000032:analysis* / Q000302:isolation & purification; D019382:Human Growth Hormone / Q000032:analysis* / Q000302:isolation & purification; D006801:Humans; D010434:Pepsin A / Q000378:metabolism; D010449:Peptide Mapping / Q000379:methods*; D021241:Spectrometry, Mass, Electrospray Ionization; D013552:Swine; D014357:Trypsin / Q000378:metabolism
D019382:Human Growth Hormone; D013006:Growth Hormone; D002918:Chymotrypsin; D014357:Trypsin; D010434:Pepsin A