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Production, partial purification, characterization, and application as laundry detergent additive of an alkalophilic protease from Bacillus velezensis strain F35
1 , 1 , 1 , 1 , 1 , 2 , * 1
1  Laboratory of Microbial Biotechnology and Engineering Enzymes (LMBEE), Centre of Biotechnology of Sfax (CBS), University of Sfax, Road of Sidi Mansour Km 6, PO Box 1177, Sfax 3018, Tunisia
2  STE JMAL (EJM)-Laundry Detergent Industry, Z.I. Avenue August 13, Z.I. Poudriere 1, P.O. Box 407, Boustene, Sfax 3000, Tunisia


Proteases represents one of the major groups of industrial enzymes and a number of detergent stable proteases have been isolated and characterized because of its widespread use in detergents. It is worthwhile to screen microbes from new habitats for proteases with novel properties to meet the needs of rapidly growing detergent industry. High-alkaline serine proteases have been successfully applied as protein degrading components of detergent formulations and are subject to extensive protein engineering efforts to improve their stability and performance. Protein engineering has been extremely used to study the structure-function relationship in proteases and led to deeper understanding of the factors influencing the cleaning performance of detergent proteases. The aim of this work is to produce an alkaline protease having characteristics allowing its potential incorporation and application in the detergence industry. Enzyme purification was conducted using the sulfate ammonium precipitation. The amplifications of the ARNr 16S and the gene encoding for the studied protease were performed by PCR. A collection of 88 strains was subjected to the screening of proteolytic activity in solid medium. Thirty-six strains showed a halo of inhibition which correspond to the protease activity. The strain F-35, isolated from wastewater at the EJM Company, which exhibited the largest halo of inhibition, was retained for further studies and assigned as Bacillus velezensis based on physiological and biochemical properties and 16S rRNA gene sequencing. The study of the effect source of carbon and azote on the activity of the studied protease showed that the best activity of 7500 U/ml was obtained by combining the Modilac (powdered milk) and the yeast extract. The characterization of the physico-chemical properties showed that the partial protease has an optimum activity at 60 °C and pH 10. The partial enzyme exhibited excellent stability and compatibility with surfactants and commercial detergents, revealing 95% stability with 2% LAS and 100% stability with Class and Arial commercial laundry detergents. The gene encoding for the studied protease was amplified by PCR and showed a size of 1.5 kb. Accordingly, such a protease could be considered as a good detergent-additive in detergent industry.

Keywords: Bacillus velezensis ; alkaline protease ; detergence formulations