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L-glutaminase is an important anticancer agent that is used extensively worldwide by depriving cancer cells of L-glutamine. The marine bacterium, Halomonas meridian was isolated from the Red Sea and selected as the more active L-glutaminase-producing bacteria. L-glutaminase fermentation was optimized at 36 h, pH 8.0, 37 °C, and 3.0% NaCl, using glucose at 1.5% and soybean meal at 2%. The purified enzyme showed a specific activity of 36.08 U/mg, and the molecular weight was found to be 57 kDa by the SDS-PAGE analysis. The enzyme was highly active at pH 8.0 and 37 °C. The kinetics’ parameters of Km and Vmax were 12.2 × 10−6 M and 121.95 μmol/mL/min, respectively, which reflects a higher affinity for its substrate. The anticancer efficiency of the enzyme showed significant toxic activity toward colorectal adenocarcinoma cells; LS 174 T (IC50 7.0 μg/mL) and HCT 116 (IC50 13.2 μg/mL). A higher incidence of cell death was observed with early apoptosis in HCT 116 than in LS 174 T, whereas late apoptosis was observed in LS 174 T more than in HCT 116. Also, the L-glutaminase induction nuclear fragmentation in HCT 116 was more than that in the LS 174T cells. This is the first report on Halomonas meridiana as an L-glutaminase producer that is used as an anti-colorectal cancer agent.

L-glutaminases are enzymes that catalyze the hydrolysis of L-glutamine, producing L-glutamate and ammonium, and they have promising applications in pharmaceutical and food industries. Several investigations have focused on thermo-tolerant L-glutaminases; however, studies on cold-adapted L-glutaminases have not been reported. These enzymes could be useful in the food industry because they display high catalytic activity at low and room temperatures, a valuable feature in processes aimed to save energy. Besides, they can be easily inactivated by warming and are suitable to prevent decomposition of thermo-labile compounds. The objectives of this work were to characterize the L-glutaminase from the Antarctic bacterium Bizionia argentinensis and analyze its capability as flavor enhancer of protein hydrolysates. The enzyme was heterologously expressed and purified from Escherichia coli, obtaining optimum and homogeneous yields. Kinetic parameters Km and Vmax were located at the lower and upper range of values reported for L-glutaminases, suggesting high catalytic efficiency. Optimum temperature was 25 °C, and the enzyme conserved around 90% of maximum activity at 0 °C and in presence of 15% (v/v) ethanol and methanol. In saline conditions, the enzyme conserved around 80% of maximum activity in 3 M NaCl. Analysis of structural model suggested cold-adaptation features such as low Arg/(Arg+Lys) ratio and fewer intramolecular interactions than mesophilic and thermo-tolerant L-glutaminases. This work provides a novel cold-adapted L-glutaminase with promising features in the food industry.

Deamination of L-glutamine to glutamic acid with the concomitant release of ammonia by the activity of L-glutaminase (L-glutamine amidohydrolase EC 3.5.1.2) is a unique reaction that also finds potential applications in different sectors ranging from therapeutics to food industry. Owing to its cost-effectiveness, rapidity, and compatibility with downstream processes, microbial production of L-glutaminase is preferred over the production by other sources. Marine microorganisms including bacteria, yeasts, and moulds have manifested remarkable capacity to produce L-glutaminase and, therefore, are considered as prospective candidates for large-scale production of this enzyme. The main focus of this article is to provide an overview of L-glutaminase producing marine microorganisms, to discuss strategies used for the lab- and large-scale production of these enzyme and to review the application of L-glutaminase from marine sources so that the future prospects can be understood.Key points• L-glutaminase has potential applications in different sectors ranging from therapeutics to food industry• Marine microorganisms are considered as prospective candidates for large-scale production of L-glutaminase• Marine microbial L-glutaminase have great potential in therapeutics and in the food industry

The aims of this study were isolation-purification and characterization of l -glutaminase from L. gasseri BRLHM clinical isolates and investigation of its efficiency as an antimicrobial agent against multidrug-resistant P. aeruginosa . The MICs of l -glutaminase and gentamicin reference were evaluated by the well-diffusion method. The biofilm on the IUD contraceptive was visualized using atomic force microscopy (AFM) image analyses. The purified l -glutaminase possessed significant antimicrobial activity against P. aeruginosa isolates ( p  < 0.05), and the antibiofilm formation activity of the purified l -glutaminase was stronger than the antibiofilm activity of the referral standard drug, gentamicin ( P  < 0.05), which were checked by the inhibition of the biofilm formation on the IUD contraceptive device. Investigations indicated that l -glutaminase may have a crucial role in future clinical applications.

l -Asparaginase is a therapeutically and industrially-competent enzyme, acting predominantly as an anti-neoplastic and anti-cancerous agent. The existing formulations of prokaryotic l -asparaginase are often toxic and contain l -glutaminase and urease residues, thereby increasing the purification steps. Production of l -glutaminase and urease free l -asparaginase is thus desired. In this research, bioprospecting of isolates from the less explored class Agaricomycetes was undertaken for l -asparaginase production. Plate assay (using phenol red and bromothymol blue dyes) was performed followed by estimation of l -asparaginase, l -glutaminase and urease activities by Nesslerization reaction for all the isolates. The isolate displaying the desired enzyme production was subjected to morphological, molecular identification, and phylogenetic analysis with statistical validation using Jukes-Cantor by Neighbour-joining tree of Maximum Likelihood statistical method. Among the isolates, Ganoderma australe GPC191 with significantly high zone index value (5.581 ± 0.045 at 120 h) and enzyme activity (1.57 ± 0.006 U/mL), devoid of l -glutaminase and urease activity was selected. The present study for the first-time reported G. australe as the potential source of l -glutaminase and urease-free l -asparaginase and also is one of the few studies contributing to the literature of G. australe in India. Hence, it can be postulated that it may find its future application in pharmaceutical and food industries.

The food enzyme glutaminase (l‐glutamine amidohydrolase; EC 3.5.1.2) is produced with the non‐genetically modified Bacillus amyloliquefaciens strain AE‐GT by Amano Enzyme Inc. The production strain met the requirements for the qualified presumption of safety (QPS) approach. The food enzyme is intended to be used in five food manufacturing processes. Dietary exposure to the food enzyme‐total organic solids (TOS) was estimated to be up to 0.462 mg TOS/kg body weight per day in European populations. Given the QPS status of the production strain and the absence of concerns resulting from the food enzyme's manufacturing process, toxicity tests were considered unnecessary by the Panel. A search for the similarity of the amino acid sequence of the food enzyme to known allergens was made and no match was found. The Panel considered that a risk of allergic reactions upon dietary exposure cannot be excluded, but the likelihood is low. Based on the data provided, the Panel concluded that this food enzyme does not give rise to safety concerns under the intended conditions of use.

Background Actinomycetes are a well-known example of a microbiological origin that may generate a wide variety of chemical structures. As excellent cell factories, these sources are able to manufacture medicines, agrochemicals, and enzymes that are crucial. Results In this study, about 34 randomly selected Streptomyces isolates were discovered in soil, sediment, sea water, and other environments. Using a qualitative fast plate assay, they were tested for L-glutaminase production, and nine of them produced a significant amount of pink L-glutamine. Streptomyces sp. strain 5 M was identified by examining the 16S rRNA gene in the promising strain G8. A pH of 7.5, an incubation temperature of 40 °C, and the use of glucose and peptone as the carbon and nitrogen sources, respectively, produced the highest quantities of L-glutaminase. The molecular weight of the isolated L-glutaminase was estimated to be 52 kDa using SDS-PAGE analysis. At pH 7.5 and Temp., 40 °C, the isolated enzyme exhibited its highest levels of stability and activity. The isolated enzyme’s K m and V max values were 2.62 mM and 10.20 U/ml, respectively. Strong toxicity against HepG-2, HeLa, and MCF-7 was observed due to the anticancer properties of the isolated L-glutaminase. Conclusion Our findings include the discovery of Streptomyces sp. strain 5 M, which yields a free L-glutaminase and maybe a possible applicant for extra pharmacological investigation as an antineoplastic drug.

Bacterial L-glutaminase (L-GLS) has emerged as a potential therapeutic target in cancer treatment by disrupting glutamine-dependent metabolic pathways in tumor cells. This study focused on isolating and characterizing L-GLS-producing marine bacteria from Mediterranean seawater for preliminary therapeutic evaluation. Halomonas aquamarina HBIM1 was identified as the most efficient isolate through comprehensive phenotypic, genotypic, and enzymatic screening. The enzyme was successfully purified, achieving a specific activity of 748.35 U/mg with 3.39-fold purification. SDS-PAGE analysis confirmed high purity with a single 66 kDa protein band. Kinetic characterization revealed optimal activity at pH 8 and 50 °C, with strong substrate affinity (Km = 0.198 mM⁻¹). Preliminary in vitro cytotoxicity screening demonstrated selective antiproliferative effects on HepG2 liver cancer cells (IC50 = 33.98 µg/ml) compared to normal WI-38 cells (IC50 = 93.43 µg/ml), yielding a 2.75-fold selectivity index. Molecular docking analysis identified tannic acid and 6-diazo-5-oxo-L-norleucine as selective inhibitors of bacterial L-GLS, with tannic acid showing the highest binding affinity (-12.25 kcal/mol) and 5-fold selectivity over human L-GLS, suggesting potential for combination therapy strategies. These proof-of-concept findings indicate the preliminary anticancer potential of Halomonas -derived L-GLS and computational support for selective inhibitor development. However, comprehensive preclinical validation, including in vivo efficacy studies, toxicological evaluation, and pharmacological profiling, is essential to establish therapeutic viability and safety before clinical consideration.

The bacterial L-asparaginase is a highly effective chemotherapeutic drug and a cornerstone of treatment protocols used for treatment the acute lymphoblastic leukemia in pediatric oncology. A potential actinomycete isolate, Streptomyces sp. strain NEAE-99, produces glutaminase-free L-asparaginase was isolated from a soil sample. This potential strain was identified as S. violaceoruber strain NEAE-99. The central composite design (CCD) approach was utilized for finding the optimal values for four variables including the mixture of soybean and wheat bran in a 1:1 ratio (w/w), the concentrations of dextrose, L-asparagine, and potassium nitrate under solid state fermentation conditions. Through the use of an artificial neural network (ANN), the production of L-asparaginase by S. violaceoruber has been investigated, validated, and predicted in comparison to CCD. It was found that the optimal predicted conditions for maximum L-asparaginase production (216.19 U/gds) were 8.46 g/250 mL Erlenmeyer flask of soybean and wheat bran mixture in a 1:1 ratio (w/w), 2.2 g/L of dextrose, 18.97 g/L of L-asparagine, and 1.34 g/L of KNO 3 . The experimental results (207.55 U/gds) closely approximated the theoretical values (216.19 U/gds), as evidenced by the validation. This suggests that the ANN exhibited a high degree of precision and predictive capability.

The food enzyme glutaminase (l‐glutamine amidohydrolase EC 3.5.1.2) is produced with the genetically modified Bacillus licheniformis strain NZYM‐JQ by Novozymes A/S. The genetic modifications do not give rise to safety concerns. The production strain met the requirements for the qualified presumption of safety (QPS). The food enzyme is free from viable cells of the production organism and its DNA. The enzyme under assessment is intended to be used in six food manufacturing processes. Dietary exposure was estimated to be up to 0.148 mg TOS/kg body weight per day in European populations. Given the QPS status of the production strain and the absence of concern resulting from the food enzyme manufacturing process, toxicological studies were not considered necessary. A search was made for the similarity of the amino acid sequence to those of known allergens and one match with a pollen allergen was found. The Panel considered that the risk of allergic reactions by dietary exposure cannot be excluded, particularly for individuals sensitised to birch and oak pollen. The Panel concluded that the food enzyme does not give rise to safety concerns under the intended conditions of use.