The cleavage site preference of the porcine pepsin on the N-terminal α1 chain of bovine type I collagen: A focal analysis with mass spectrometry

Abstract

Bovine type I collagen consists of two α1 and one α2 chains, containing the internal triple helical regions and the N- and C-terminal telopeptides. In industries, it is frequently digested with porcine pepsin to produce a triple helical collagen without the telopeptides. However, the digestion mechanism is not precisely understood. Here, we performed a mass spectrometric analysis of the pepsin digest of the N-terminal telopeptide pQLSYGYDEKST- GISVP (1-16) in the α1 chain. When purified collagen was digested, pQLSYGY (1-6) and pQLSY- GYDEKSTG (1-12) were identified, while DEKSTG (7-12) was not. When the N-terminal telopeptide mimetic synthetic peptide pQLSK(MOCAc) GYDEKSTGISK(Dnp)P-NHi2 was digested, pQLSK (MOCAc)GYDEKSTG (1-12) and ISK(Dnp)P-NH2(13-16) were readily identified, pQLSK(MOCAc) GY (1-6) and DEKSTGISK(Dnp)P-NH2 (7-16) were weakly detected, and DEKSTG (7-12) was hardly identified. These results suggest that pepsin preferentially cleaves Tyr6-Asp7 and less preferentially Gly12-Ile13. They also suggest that the former cleavage requires native collagen structure, while the latter cleavage does not.