Recombinant production and characterization of L-glutaminase

Because of the therapeutical impacts of hydrolytic enzymes in different diseases, in particular malignancies, we aimed to produce a recombinant putative L-glutaminase (GLS A.sub.SL-1) from a recently characterized halo-thermotolerant Bacillus sp. SL-1. For this purpose, the glsA gene was identified...

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Bibliographic Details
Published in:Applied microbiology and biotechnology 2022-09, Vol.106 (17), p.5511
Main Authors: Simay, Shayan, Akbarzadeh-Khiavi, Mostafa, Pourseif, Mohammad M, Barar, Jaleh, Safary, Azam, Omidi, Yadollah
Format: Article
Language:English
Subjects:
Physiological aspects
Recombinant proteins
Online Access:Get full text
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Summary:Because of the therapeutical impacts of hydrolytic enzymes in different diseases, in particular malignancies, we aimed to produce a recombinant putative L-glutaminase (GLS A.sub.SL-1) from a recently characterized halo-thermotolerant Bacillus sp. SL-1. For this purpose, the glsA gene was identified and efficiently overexpressed in the Origami[TM] B (DE3) strain. The yield of the purified GLS A.sub.SL-1 was ~ 20 mg/L, indicating a significant expression of recombinant enzyme in the Origami. The enzyme activity assay revealed a significant hydrolytic effect of the recombinant GLS A.sub.SL-1 on L-asparagine (Asn) (i.e., K.sub.m 39.8 [mu]M, k.sub.cat 19.9 S.sup.-1) with a minimal affinity for L-glutamine (Gln). The GLS A.sub.SL-1 significantly suppressed the growth of leukemic Jurkat cells through apoptosis induction (47.5%) in the IC.sub.50 dosage of the enzyme. The GLS A.sub.SL-1 could also change the Bax/Bcl2 expression ratio, indicating its apoptotic effect on cancer cells. The in silico analysis was conducted to predict structural features related to the histidine-tag exposure in the N- or C-terminal of the recombinant GLS A.sub.SL-1. In addition, molecular docking simulation for substrate specificity revealed a greater binding affinity of Asn to the enzyme binding-site residues than Gln, which was confirmed in experimental procedures as well. In conclusion, the current study introduced a recombinant GLS A.sub.SL-1 with unique functional and structural features, highlighting its potential pharmaceutical and medical importance. GLS A.sub.SL-1 represents the first annotated enzyme from Bacillus with prominent asparaginase activity, which can be considered for developing alternative enzymes in therapeutic applications.
ISSN:0175-7598
DOI:10.1007/s00253-022-12058-y