title: Collagen V Is a Potential Substrate for Clostridial Collagenase G in Pancreatic Islet Isolation.
abstract: The clostridial collagenases, H and G, play key roles in pancreatic islet isolation. Collagenases digest the peptide bond between Yaa and the subsequent Gly in Gly-Xaa-Yaa repeats. To fully understand the pancreatic islet isolation process, identification of the collagenase substrates in the tissue is very important. Although collagen types I and III were reported as possible substrates for collagenase H, the substrate for collagenase G remains unknown. In this study, collagen type V was focused upon as the target for collagenases. In vitro digestion experiments for collagen type V were performed and analyzed by SDS-PAGE and mass spectrometry. Porcine pancreatic tissues were digested in vitro under three conditions and observed during digestion. The results revealed that collagen type V was only digested by collagenase G and that the digestion was initiated from the N-terminal part. Tissue degradation during porcine islet isolation was only observed in the presence of both collagenases H and G. These findings suggest that collagen type V is one of the substrates for collagenase G. The enzymatic activity of collagenase G appears to be more important for pancreatic islet isolation in large mammals such as pigs and humans.
authors.affiliation: Department of Biochemistry, Tohoku University Graduate School of Medicine, Sendai 980-8575, Japan; AMED-CREST, Japan Agency for Medical Research and Development, Tokyo 100-0004, Japan.
authors.forename: Hiroki
authors.initials: H
authors.lastname: Shima
authors.affiliation: Division of Transplantation and Regenerative Medicine, Tohoku University School of Medicine, Sendai 980-8575, Japan.
authors.forename: Akiko
authors.initials: A
authors.lastname: Inagaki
authors.affiliation: Division of Transplantation and Regenerative Medicine, Tohoku University School of Medicine, Sendai 980-8575, Japan; New Industry Creation Hatchery Center, Tohoku University, Sendai 980-8579, Japan.
authors.forename: Takehiro
authors.initials: T
authors.lastname: Imura
authors.affiliation: Graduate School of Agricultural Science, Tokyo University of Agriculture and Technology, Fuchu 183-8509, Japan.
authors.forename: Youhei
authors.initials: Y
authors.lastname: Yamagata
authors.affiliation: New Industry Creation Hatchery Center, Tohoku University, Sendai 980-8579, Japan.
authors.forename: Kimiko
authors.initials: K
authors.lastname: Watanabe
authors.affiliation: Department of Biochemistry, Tohoku University Graduate School of Medicine, Sendai 980-8575, Japan; AMED-CREST, Japan Agency for Medical Research and Development, Tokyo 100-0004, Japan; Center for Regulatory Epigenome and Diseases, Tohoku University Graduate School of Medicine, Sendai 980-8575, Japan.
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Igarashi
authors.affiliation: Division of Transplantation and Regenerative Medicine, Tohoku University School of Medicine, Sendai 980-8575, Japan.
authors.forename: Masafumi
authors.initials: M
authors.lastname: Goto
authors.affiliation: Graduate School of Biomedical Engineering, Tohoku University, Sendai 980-8575, Japan.
authors.forename: Kazutaka
authors.initials: K
authors.lastname: Murayama
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000494:pharmacology*; D003013:Clostridium / Q000201:enzymology; D024062:Collagen Type V / Q000187:drug effects* / Q000378:metabolism; D017364:Collagenases / Q000494:pharmacology*; D004591:Electrophoresis, Polyacrylamide Gel; D066298:In Vitro Techniques; D007515:Islets of Langerhans / Q000187:drug effects* / Q000378:metabolism; D016381:Islets of Langerhans Transplantation; D013058:Mass Spectrometry; D003012:Microbial Collagenase / Q000494:pharmacology*; D013552:Swine
chemical_list: D001426:Bacterial Proteins; D024062:Collagen Type V; C118796:ColH protein, Clostridium histolyticum; D017364:Collagenases; D003012:Microbial Collagenase