Biochemical Comparison Between Clostridium Hystoliticum Collagenases G and H obtained by DNA Recombinant and Extractive Procedures

Abstract

One of the main success practices in cell therapy is the transplantation of allogeneic pancreatic islets cells in patients with type 1 diabetes. However, at the moment less than 50 % of procedures are successfully completed; this is because the procedure of pancreatic islet isolation/purification, are not well standardized. This low percent of success can be ascribed to several factors, including: - the variability in donor characteristics, - pancreas recovery and preservation, - not the last, an unpredictable enzymatic blend efficiency. At present, the process of pancreatic islets isolation, sees the use of proteolytic enzymes, collagenases (Class I and Class II) produced by Clostridium hystoliticum bacteria, and purified by extractive procedures. Limit of these enzymes lies in composition variability, such as their concentration and the presence of toxic components, together autocatalytic processes and the presence of other proteolytic enzymes. To reduce the variability in collagenase composition in the blends usable in cell isolation, and to minimize the variability in enzyme composition, we produced the major classes of C. hystoliticum collagenases: collagenase G and collagenase H by DNA recombination techniques. Using biochemical/ezymologic approaches, we compared composition, enzymatic activity and auto-digestion processes, of the C. hystoliticum collagenases obtained using extraction procedures versus the recombinant ones.