Mutation of AN39-1 for production and characterization of constitutive, thermostable and pH-resistant dextransucrase

Abstract

Objective: Leuconostoc mesenteroides AN39-1 has recently been isolated from Crataegus orientalis var. Orientalis. It produces inducible extracellular dextransucrase (EC 2.4.1.5) forming dextran from sucrose. The aim of this study was (1) to obtain constitutive, pH-resistant and thermostable dextransucrase, (2) to characterization of these dextransucrase. Methods: Mutagenesis was carried out on the parent strain (AN39-1) using UV, ethyl methane sulfonate, and N-methyl-N'-nitro-N-nitrosoguanidine. Dextransucrases from wild type (AN39-1) and the mutant strain (A26-2/11) were purified by polyethylene glycol (PEG) precipitation and characterized. Results: Mutants (A26, A26-2, and A26-2/11) hyper producing and constitutive for dextransucrase were isolated. The mutants (A26, A26-2, A26-2/11) produced 7.2, 8.1, and 2.0 times more dextransucrase activity as compared to parent strain on sucrose medium, respectively. In addition, the mutants produced dextransucrase on glucose medium with higher activities (3.0-5.8 times) than what the parental strain produced on sucrose medium. The mutant enzyme (A26-2/11) was much more thermostable than the native enzyme and resistant to pH more than dextransucrase of AN39-1. The dextransucrase from mutant strain was stable up to 35 degrees C and pH of 7.5 for 3 hr. Conclusion: The structures of dextrans produced by wild type and mutant enzymes were similar to commercially produced B-512 F dextran. Thus, the newly dextransucrases produced by mutant strain could find industrial applications at higher temperature and pH.