Cloning, Expression and Characterization of Thermophilic and Alkalophilic N-acetylglucosaminidase from Streptomyces sp NK52 for the Targeted Production of N-acetylglucosamine

A chitinolytic Streptomyces strain isolated from an alkaline habitat produced six different isozymes of chitinases. PCR amplification of DNA with chitinase domain specific primers yielded six amplicons of which, a 0.8 Kb fragment was cloned in pQE-30UA direct cloning vector and transformed into E. coli M15 cells (pREP4). The recombinant homodimer protein had a molecular mass of 44 kDa, and the 22 kDa monomers displayed 45 and 60 % activity in the presence of reducing agents. The size of the monomers is close to the predicted putative ORF of 17.8 kDa. The enzyme exhibits extreme pH and temperature optima of 10.0 and 70 A degrees C respectively making it favorable for industrial applications. Its gene sequence revealed no homology with the reported N-acetylglucosaminidases suggesting that it could have novel attributes. This enzyme could be useful in the large scale production of N-acetylglucosamine which is currently having numerous therapeutic and commercial applications.