L-Asparaginase Synthesis by some Egyptian Bacterial Isolates and Molecular Identification Based on the 16S rRNA Sequence

Khadiga A. Aboutaleb and Naglaa Mohammed Ebeed ; Abou-Taleb, Khadiga 


Abstract


The increasing importance of L-Asparaginase (LAse) in recent years for its therapeutic applications as well as extensive uses in food industries prompted to utilize newer microbial sources for L-Asparaginase production. The present study aimed to isolate microbial species that produce LAse. In addition to, molecular identification of the most potent isolate for LAse enzyme activity. Sixty isolates of bacteria were isolated from various plant rhizosphere soils and were screened for their ability to produce L-asparaginase on modified M-9 solid medium containing asparagine as nitrogen source and phenol red as an indicator. The diameter of the produced pink (red) zone ranged between 0.01 and 2.7 cm. Four isolates; PS14, SF2, SO5 and SW5 showed the highest L-asparaginase activity among the isolates were selected. Nutritional and cultural conditions affecting the production of L-asparaginase by the tested isolates were optimized. Maximal production of L-asparaginase was recorded on the presence of sucrose 2.0 g/l and yeast extract 0.50% plus L-asparagine 0.50% in the production medium served as good carbon and nitrogen sources, respectively, and pH was adjusted to 7.5. The incubation temperature at 30°C and shaking speed at 150 rpm gave the highest yield of L-asparaginase after 48 h of incubation period for tested isolates using shake flasks as a batch culture. The enzyme yielding capacity from isolates PS14, SF2, SO5 and SW5 in the optimized medium were found to be 1.85 and 1.45, 1.26 and 1.21 folds higher than unoptimized medium, respectively. The potent 4 isolates were differentiated based on nucleotide sequences of 16S rDNA gene. Phylogenetic analysis based on 16S rDNA gene sequence revealed that these isolates belong to the genus Acinetobacter and genus Bacillus. The partial 16S rDNA gene sequences obtained in the present study were deposited in GenBank database under accession number KM235291, KM266616, KM266617 and KM266618 for PS14, SF2, SO5 and SW5 strains respectively.


Other data

Keywords L-asparaginase activity, 16S rDNA analysis, bacteria, nutritional factors, cultural conditions
Issue Date 2013
Source Aboutaleb, Kh. A. & Ebeed, N. M. (2013). L-Asparaginase Synthesis by some Egyptian Bacterial Isolates and Molecular Identification Based on the 16S rRNA Sequence. Middle East Journal of Applied Sciences, 3(4): 231-243.
Journal Middle East Journal of Applied Sciences 
URI http://research.asu.edu.eg/handle/123456789/2309


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