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

• Published in: Bioscience, biotechnology, and biochemistry biotechnology, and biochemistry, 2017-03, Vol.81 (3), p.514-522
• Main Authors: Qian, Jun, Ito, Shinji, Satoh, Junko, Geng, Hongmin, Tanaka, Keisuke, Hattori, Shunji, Kojima, Kenji, Takita, Teisuke, Yasukawa, Kiyoshi
• Format: Article
• Language: English
• Subjects: Animals; Cattle; collagen; Collagen Type I - chemistry; Collagen Type I - isolation & purification; Collagen Type I - metabolism; Electrophoresis, Polyacrylamide Gel; Fluorescence; mass spectrometry; pepsin; Pepsin A - chemistry; Pepsin A - metabolism; Peptide Fragments - chemistry; Skin - chemistry; Sus scrofa; Tandem Mass Spectrometry - methods; telopeptide
• ISSN: 0916-8451; 1347-6947
• DOI: 10.1080/09168451.2016.1263146

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 pQLSYGYDEKSTGISVP (1-16) in the α1 chain. When purified collagen was digested, pQLSYGY (1-6) and pQLSYGYDEKSTG (1-12) were identified, while DEKSTG (7-12) was not. When the N-terminal telopeptide mimetic synthetic peptide pQLSK(MOCAc)GYDEKSTGISK(Dnp)P-NH 2 was digested, pQLSK(MOCAc)GYDEKSTG (1-12) and ISK(Dnp)P-NH 2 (13−16) were readily identified, pQLSK(MOCAc)GY (1−6) and DEKSTGISK(Dnp)P-NH 2 (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. Pepsin preferentially cleaves Tyr6-Asp7 and less preferentially Gly12-Ile13. The former cleavage requires native collagen structure, while the latter cleavage does not.