Purification and characterization of a carboxymethyl cellulase from Artemia salina

Abstract

The first process in ethanol production from marine macroalgae is the breakdown of polymers. These days, this process is accomplished by heating raw materials in acidic condition which is energy consuming and environmentally unfriendly. Brine shrimp (Artemia salina) belonging to a group of crustaceans that feed on microalgae, which require a cellulase enzyme that may be used in ethanol production from marine algae. Protein that has possibility with cellulase activity was purified and the activity was analyzed in different condition. The Congo red overlay method was done to distinguish whether A. salina is the real source of the cellulase enzyme. PCR using 16s rRNA primers was also done to confirm that there was no cellulase producing bacterial contamination. Ammonium sulfate fractionation, gel filtration and ion exchange chromatography were performed sequentially, for the purification of the novel cellulase. SDS-PAGE results revealed that the cellulase enzyme has a molecular mass of ~96 kDa. Temperature, pH and salinity values were found to be optimum at 55°C, pH8.0 and 600 mM NaCl, respectively. Additionally, purified proteins were processed through UPLC and mass spectrophotometry. Based on the analyzed data using ProteinLynx Global Server (PLGS), one of the protein sequences revealed cellulase protein (endo-beta-1, 4-glucanase). As cellulases are biotechnologically significant inducible enzymes, challenges on their commercial production and efficient utilization arise. Adaptation in an environment with high salinity could contribute for the utilization of this enzyme in the future in the field of bio-energy processes. Hence, enhancement in the process efficiency by searching for more active cellulases has been of great scientific interest.