BIOTECHNOLOGICALLY RELEVANT ENZYMES AND PROTEINS Characterisation of a detergent-stable alkaline protease from a novel thermophilic strain Paenibacillus tezpurensis sp. nov. AS-S24-II Sudhir K. Rai & Jetendra K. Roy & Ashis K. Mukherjee Received: 25 June 2009 / Revised: 25 June 2009 / Accepted: 13 July 2009 / Published online: 11 August 2009 # Springer-Verlag 2009 Abstract An alkaline-protease-producing bacterial strain (AS-S24-II) isolated from a soil sample in Assam is a Gram-stain-positive, catalase-positive, endospore-forming rod and grows at temperatures ranging from 30 °C to 60 °C and salinity ranging from 0% to 7% (w/v) NaCl. Phenotypic characterisation, chemotaxonomic properties, presence of Paenibacillus-specific signature sequences, and ribotyping data suggested that the strain AS-S24-II represents a novel species of the genus Paenibacillus, for which the name Paenibacillus tezpurensis sp. nov. (MTCC 8959) is proposed. Phylogenetic analysis revealed that P. lentimorbus strain DNG-14 and P. lentimorbus strain DNG-16 represent the closest phylogenetic neighbour of this novel strain. Alkaline protease production (598×10 3 Ul -1 ) by P. tezpurensis sp. nov. in SmF was optimised by response surface method. A laundry-detergent-stable, Ca 2+ -independent, 43-kDa molecular weight alkaline serine protease from this strain was purified with a 1.7- fold increase in specific activity. The purified protease displayed optimum activity at pH 9.5 and 45–50 °C temperature range and exhibited a significant stability and compatibility with surfactants and most of the tested commercial laundry detergents at room temperature. Further, the protease improved the wash performance of detergents, thus demonstrating its feasibility for inclusion in laundry detergent formulations. Keywords Alkaline protease . Detergent compatibility . Paenibacillus tezpurensis . Response surface . Serine protease . Submerged fermentation Introduction Microbial proteases, representing one of the three largest groups of industrial enzymes, account for approximately 60% of the total enzyme sale. Amongst the different types of proteases, alkaline proteases particularly those isolated from the genus Bacillus find a wide range of applications in laundry, dishwashing, textile, food processing, pharmaceuticals, leather, paper and pulp industries (Anwar and Saleemuddin 1998; Gupta et al. 2002). Owing to the better cleansing properties of enzyme-based detergents at lower washing temperature and pollution-alleviating capacity over conventional syn- thetic detergents (Krik et al. 2002; Mukherjee et al. 2008), alkaline proteases have made their way as key ingredients in detergent formulations (Maurer 2004). It is well known that exploration of microbial diversity is the major driving force for the development of biotechno- logical products and processes. The northeastern part of India is considered as one of the mega biodiversity zones of the world and, by screening the soil samples of this region, we have isolated a potent bacterial strain capable of secreting significant amount of protease in the culture medium at 50 °C temperature. Taxonomic identification of the strain by polyphasic approach showed that this is a novel species belong to the genus Paenibacillus, and this strain was subsequently named as Paenibacillus tezpurensis sp. nov. There are hardly few reports available on protease production by Paenibacillus which are ubiquitous in nature, and the proteases from this genus may find novel industrial The 16S rDNA sequence to GenBank and the accession number are bankit1191281 FJ804507, and the strain is deposited in Microbial Type Culture Collection (MTCC 8959). S. K. Rai : J. K. Roy : A. K. Mukherjee (*) Department of Molecular Biology and Biotechnology, School of Science and Technology, Tezpur University, Tezpur 784 028 Assam, India e-mail: akm@tezu.ernet.in Appl Microbiol Biotechnol (2010) 85:1437–1450 DOI 10.1007/s00253-009-2145-y