Alkali-treated collagen retained the triple helical conformation and the ligand activity for the cell adhesion via alpha2beta1 integrin.
Alkaline treatment is a good method for extracting collagen with high recovery even from an aged animal specimen. However, the properties of collagen treated under alkaline conditions have not been well established yet. By the treatment with a solution of 3% sodium hydroxide and 1.9% monomethylamine, the isoelectric point of type I collagen was lowered from 9.3 to 4.8 because of the conversions of Asn and Gln to Asp and Glu. With the acidification of the pI, the denaturation temperature of the collagen was decreased from 42 to 35 degrees C after 20 d treatment, but the collagen-specific triple helical conformation was maintained. Human keratinocytes and fibroblasts adhered to the alkali-treated collagen via the collagen receptor integrin alpha2beta1. This indicates that the alkali-treated collagen maintained its property as a biological adherent molecule. Unlike acid-soluble collagen, alkali-treated collagen lost the ability to form fibrils at neutral pH under physiological conditions. This ability was lost even after 4 h of alkaline treatment, when the denaturation temperature of the collagen did not change. On the other hand, the alkali-treated collagen formed a fibrous precipitate with a uniform diameter of 50-70 nm under acidic conditions at 30 degrees C.
Hattori
S
S
Nippi Research Institute of Biomatrix, Adachi-ku, Tokyo, 120-8601, Japan. shunhatt@bekkoame.ne.jp
Adachi
E
E
Ebihara
T
T
Shirai
T
T
Someki
I
I
Irie
S
S
D000468:Alkalies; D000595:Amino Acid Sequence; D000818:Animals; D002417:Cattle; D002448:Cell Adhesion / Q000502:physiology; D002478:Cells, Cultured; D003094:Collagen / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D016023:Integrins / Q000378:metabolism; D007526:Isoelectric Point; D015603:Keratinocytes / Q000166:cytology / Q000378:metabolism; D008024:Ligands; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D011487:Protein Conformation; D011489:Protein Denaturation; D038981:Receptors, Collagen; D012867:Skin / Q000737:chemistry
D000468:Alkalies; D016023:Integrins; D008024:Ligands; D038981:Receptors, Collagen; D003094:Collagen