Research Topic:This research intends to characterize the major proteases secreted by two distinct morphologies of the marine bacterium Teredinobacter turnirae. In the course of laboratorial experiments, it was found that this microorganism is pleomorphic, capable of shanging its morphology from a rod-like (background of this page) to a clump-like appearance. Both morphologies secrete proteolytic activity, with clumps being able to produce about 20 - 30 times more as compared with rods.

    Proteases have principally been used as detergent additives, preparing tannable skins and recovering hair from animal pelts, cleaning raw silk, silver recovery from film, brewing and gelatin manufacture. They account for 40% of the enzyme market, currently estimated to have the value of 1.4 billion dollars.

    One protease was previously isolated and characterized. The enzyme was uniquely characterized by its stability over a broad pH range as well as a tolerance under a variety of chemical environments. The stability and nearly complete retention of activity in high alkaline, saline and oxidizing environments of this product exceeds that of commercially available proteases. Since most commercial uses of alkaline proteases involve one or more of these conditions, this represents a clear advantage and a potential opportunity to exploit an sixty million dollar market available to alkaline proteases. This alkaline protease is effective in non-phosphate detergents that are being increasingly mandated to decrease the level of phosphate pollution in lakes and streams. Thus, the product can potentially contribute to decrease environmental impact of detergent use.

    The project was approached by initially studying the microbiological and biochemical characteristics of the marine bacterium (carbon and nitrogen sources for best growth and production of proteolytic activity). Upon finding the different morphology, growth studies were implemented, from which it was concluded that the clump cells showed differences in their requirements for media components, metabolism and, more interestingly, in the capability of growing in media with no added NaCl. With so many differences being observed, the question of the nature of the enzyme responsible for the high readings associated with the clump morphology arised.

    It was found that the major protease associated with the rod cells (rods protease) was related, yet different, to that previoulsy isolated by other researchers: values for molecular weight, specific activity, stability and activity for pH, temperature and ionic strength, and sensitivity to inhibitors were similar. The major protease isolated from the clump cells (clumps protease) had a specific activity that was 50-fold higher than that isolated for the rods, which explains partially the increased titers measured in the broth of these cells. Additionally, the protease was very different from the rods protease in its sensitivity to protease inhibitors. Finally, a N-sequence analysis was done to compare the enzymes.

    It was concluded that the metabolic and morphological differences observed for the rods and clumps were accompanied by a difference in the major protease being secreted. The protease associated with the clump-cells was more versatile (trypsin, chymotrypsin and subtilisin activities) while rods protease showed to be specific towards trypsin-like substrates.

    This project is supported by the United States Department of Agriculture and Praxis XXI.
 

    (The background of this page is an original picture of the marine bacterium Teredinobacter turnirae)