Comparative structural and kinetic study for development of a novel candidate L-asparaginase based pharmaceutical

Abstract

L-asparaginases (L-ASNase, EC 3.5.1.1) are amidohydrolases catalysing the conversion of L-Asn to L-Asp and to a lower extent L-Gln to L-Glu. This enzyme is used for the treatment of acute lymphoblastic leukemia. Currently, Escherichia coli and Erwinia chrysanthemiL-ASNases are used as oncological drugs. However, harmful side-effects and hypersensitivity reactions are the main limitations of these therapeutics. A link has been proposed between L-GLNase activity and harmful side effects. Therefore, finding new L-ASNases with low L-GLNase activity is important for medical applications. A detailed biochemical characterization of a wide range of L-ASNases linked with in-silico approaches could contribute to discovering better oncologic L-ASNase candidates. In this study, ten bacterial and yeast l-ASNase belonging to type I and II classes of l-ASNase families were characterized. The optimum pH and temperature of the l-ASNases were at pH 7.0–9.0 and 35–50 °C range, respectively. None of the ten l-ASNases displayed detectable l-GLNase activity. Structural comparisons of these ten L-ASNases with quite differing kinetic properties showed that the residues with a catalytic role are conserved and some differences at position 59 close to the substrate may affect the kinetic parameters. The type I l-ASNase from Lachancea thermotolerans yeast (LtASNase) exhibited the highest specific activity (313.82 U/mg) and catalytic efficiency for l-Asn. Therefore, LtASNase is a promising candidate oncological therapeutic for further investigation in pharmaceutical applications.