From transcriptome to cloning, expression, purification, biochemical characterization and biotechnological application of a serine protease S8 with collagenolytic activity

• Grant number: 25/05734-0
• Support Opportunities: Scholarships in Brazil - Doctorate
• Start date: June 01, 2025
• End date: February 28, 2027
• Field of knowledge: Biological Sciences - Microbiology - Applied Microbiology
• Principal Investigator: Hamilton Cabral
• Grantee: Camila Langer Marciano
• Host Institution: Faculdade de Ciências Farmacêuticas de Ribeirão Preto (FCFRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil
• Associated research grant: 24/18725-6 - From transcriptome to cloning, expression, purification, biochemical characterization and biotechnological application of a serine protease S8 with collagenolytic activity, AP.R

Abstract

Since ancient times, microorganisms have been widely used to obtain various products of human interest. With technological advances, fermentation bioprocesses have become essential for the large-scale production of metabolites, including proteins and enzymes. Among these enzymes, proteases play crucial roles in physiological and industrial processes, and a specific group of them, collagenases, have a number of applications due to their ability to degrade collagen, especially in the health area, where they are used in the enzymatic debridement of chronic wounds. Currently, Clostridium sp. is the main source of collagenase for industrial application; however, the pathogenic and anaerobic nature of this microorganism limits the application of the enzyme. Thus, there is a growing demand for alternative sources of non-pathogenic microorganisms or for recombinant methods that enable the safe and economical production of this enzyme. In this context, the fungus Purpureocilium lilacinum, found in an environment of decomposing organic matter, has unexplored potential for the production of collagenases and other hydrolases. A study by our research group with the wild fungus identified the potential of two secreted proteases with collagenolytic activity. The transcriptomic study, also carried out by our research group, identified a serine protease of the S8 subfamily, which, based on in silico analyses, demonstrates collagenolytic potential. In addition to the known applications of collagenases, collagen hydrolysis can be explored to obtain bioactive peptides, being used in several sectors. Thus, the study and application of collagenolytic proteases emerge as promising areas in biotechnology and regenerative medicine. Therefore, the objective of this project is to obtain recombinant serine protease S8, purify, characterize and develop an ointment with healing properties, formulated from recombinant collagenase, in addition to evaluating the biological activities of the peptides obtained by the action of the enzyme on collagen and gelatin due to their possible potential for biotechnological applications.