The role of extraction method to collagen substrates in enzymolysis of type I collagenase.

Collagens are ubiquitous biomaterials in animal tissues whose characteristic triple-helical structure can only be hydrolyzed under physiological conditions by a few specific proteases. At present, information on the differences of collagenase hydrolysis behavior to collagen substrate caused by extraction methods is still lacking. Acid-relaxed extracted collagen (ARC) and acetic acid-pepsin extracted collagen (APC) were obtained from bovine hide by acetic acid and acetic acid-pepsin extraction method, respectively. The enzymolysis behavior of type I collagenase on ARC and APC were investigated by means of fluorescence spectra, UV spectra, and determination the release of hydrolysates into the supernatant. The results revealed that APC showed a lower molecular weight, a higher pI (5.59) and denaturation temperature (T = 66.9 °C) than that of ARC (pI = 4.67, T = 57.8 °C). Moreover, APC demonstrated greater resistance to type I collagenase than ARC. The cleavage on the non-helical terminal domains by pepsin might play the role in the better thermal stability, the higher pI, and the more collagenase resistance of APC than ARC. The findings of this work should provide new insights into collagenase hydrolysis behavior and facilitate targeted utilization of collagen extracted by various method.