title: Novel glutaminase free L-asparaginase from Nocardiopsis alba NIOT-VKMA08: production, optimization, functional and molecular characterization.
abstract: Studies were carried out for the optimization and production of novel extracellular glutaminase-free L-asparaginase from Nocardiopsis alba NIOT-VKMA08. Among the tested carbon and nitrogen sources, maximum L-asparaginase production was observed with a combination of L-asparagine and maltose (1.5%) and twofold increase in yield (18.47 IU mL(-1)) was observed with newly optimized NIOT-asparaginase medium. Activity of the purified enzyme was moderately inhibited by various divalent cations and thiol group blocking reagents, with K(m) and V(max) of 0.127 mM and 5.50 U µg(-1). Optimum pH and temperature of purified L-asparaginase for the hydrolysis of L-asparagine was 8.0 and 37 °C, respectively. The enzyme inhibited polyacrylamide formation in 10% solution and it was very specific for its natural substrate L-asparagine. Partial glutaminase activity was not detected, which could reduce the possibility of side effects during cancer therapy. L-Asparaginase biosynthesis gene (ansA) was cloned and transformed in E. coli JM109. The ansA gene sequence reported in this study contains several base substitutions with that of reported sequences in GenBank, resulting in altered amino acid sequences of the translated protein.
authors.affiliation: Andaman and Nicobar Centre for Ocean Science and Technology, Earth System Sciences Organization, National Institute of Ocean Technology (ESSO-NIOT), Dollygunj, Port Blair, 744 103, Andaman and Nicobar Islands, India, bmeena79@yahoo.com.
authors.forename: Balakrishnan
authors.initials: B
authors.lastname: Meena
authors.forename: Lawrance
authors.initials: L
authors.lastname: Anburajan
authors.forename: Palaiya Sukumaran
authors.initials: PS
authors.lastname: Dheenan
authors.forename: Mehmuna
authors.initials: M
authors.lastname: Begum
authors.forename: Nambali Valsalan
authors.initials: NV
authors.lastname: Vinithkumar
authors.forename: Gopal
authors.initials: G
authors.lastname: Dharani
authors.forename: Ramalingam
authors.initials: R
authors.lastname: Kirubagaran
mesh_terms: D039903:Actinobacteria / Q000145:classification* / Q000201:enzymology*; D000595:Amino Acid Sequence; D001215:Asparaginase / Q000096:biosynthesis* / Q000737:chemistry* / Q000302:isolation & purification; D004789:Enzyme Activation; D004795:Enzyme Stability; D005972:Glutaminase / Q000737:chemistry*; D008969:Molecular Sequence Data; D013045:Species Specificity; D013379:Substrate Specificity
chemical_list: D001215:Asparaginase; D005972:Glutaminase