Comparative Biochemical Characterization of L-Asparaginases from Four Species of Lactic Acid Bacteria

Abstract

L-Asparaginase (ASNase; EC 3.5.1.1) is an enzyme that catalyzes the hydrolysis of L-asparagine to L-aspartic acid and ammonia. Generally, ASNases from Escherichia coli and Erwinia chrysanthemi are used for the treatment of acute lymphoblastic leukemia. However, few studies focusing on ASNase from lactic acid bacteria (LAB) have been reported. The aim of this study is to characterize ASNase genes from four LAB strains: Streptococcus thermophiles, Lactobacillus plantarum, L. acidophilus, and L. sakei. ASNase genes from each strain amplified by polymerase chain reaction PCR were inserted into NdeI and XhoI sites of pET28a-(+) and cloned in E. coli BL21(DE3). Recombinant ASNases were purified using nickel-nitrilotriacetic acid column chromatography. Among the four strains, the purified recombinant ASNase from S. thermophilus exhibited the highest specific activity of 113.0 U/mg and specificity for L-asparagine. The pH and temperature ranges for S. thermophilus ASNase were pH 8.0-9.0 and 30°C-50°C, respectively. The activity of the enzyme was significantly inhibited by Ni 2+ . K m and k cat values were 2.91 mM and 1.53 × 10 2 s -1 , respectively. In this study, we described the biochemical properties of ASNases from four LAB and demonstrated that ASNase from S. thermophilus has potential applications in food processing.

Metagenomic discovery and functional validation of L-asparaginases with anti-leukemic effect from the Caspian Sea

“…The speci c activities of the puri ed enzymes were 700, 240 and 100 U/mg respectively for CAspI, CAspII and CAspIII while the speci c activity of the commercial L-asparaginases from E. coli and E. chrysanthemi are between 280-400 U/mg and 650-700 U/mg respectively [4]. Phetsri et al, reported maximum speci c activity of 113 U/mg for Streptococcus thermophiles among four species of lactic acid bacteria tested [52]. Speci c activities of 833 and 155 U/mg are also stated for the L-asparaginases puri ed from Thermococcus Kodakarensis and Acinetobacter soli, respectively [53,54].…”

Metagenomic discovery and functional validation of L-asparaginases with anti-leukemic effect from the Caspian Sea

“…The specific activities of the purified enzymes were 700, 240, and 100 U/mg, respectively, for CAspI, CAspII, and CAspIII while the specific activity of the commercial L-asparaginases from E. coli and E. chrysanthemi is between 280 and 400 U/mg and 650-700 U/mg, respectively (Narta et al, 2007). Phetsri et al reported maximum specific activity of 113 U/mg for Streptococcus thermophiles among four species of lactic acid bacteria tested (Phetsri et al, 2019). Specific activities of 833 and 155 U/mg are also stated for the L-asparaginases purified from Thermococcus kodakarensis and Acinetobacter soli, respectively (Jiao et al, 2020;Chohan et al, 2020).…”