Comparison of Enzymatic Activity of Novel Recombinant L-asparaginases of Extremophiles

Думина, М. В.; Zhgun, A. A.; Pokrovskay, M. V.; Александрова, С. С.; Zhdanov, Dmitry; Соколов, Н. Н.; Eldarov, Michael A.

doi:10.1134/s0003683821050057

Cited by 9 publications

(6 citation statements)

References 39 publications

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“…However, this classification has been disputed due to the absence of the thermophilic group. According to Dumina et al ( Dumina et al, 2021 ), thermophilic L-ASNases differ from mesophiles both in their structural properties (they can be found in a hexameric form ( Pritsa and Kyriakidis, 2001 )), topological properties, deviation in the canonical arrangement of their active site, etc. For this reason, a new classification for thermophilic L-ASNases (class IV) proposed by the authors would be included in this review ( Figure 3 ).…”

Section: Type and Characteristics Of L-asnase In Organismsmentioning

confidence: 99%

“…The L-ASNase were expressed in E. coli , and their codon composition was optimized using the “Twist Codon Optimization” (Twist Bioscience, USA) tool ( TwistBioscience, 2022 ). This strategy allowed to obtain activities of 0.6 U/mL at 24°C and pH 9.6 for S. borealis ; 2.6 U/mL at 94°C and pH 5.2 for A. saccharovorans ; and 9.6 U/mL at 37°C and pH 9.6 for M. roseu ( Dumina et al, 2021 ).…”

Section: Improvement In Systems For L-asnase Heterologous Expressionmentioning

confidence: 99%

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Current state of molecular and metabolic strategies for the improvement of L-asparaginase expression in heterologous systems

et al. 2023

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Heterologous expression of L-asparaginase (L-ASNase) has become an important area of research due to its clinical and food industry applications. This review provides a comprehensive overview of the molecular and metabolic strategies that can be used to optimize the expression of L-ASNase in heterologous systems. This article describes various approaches that have been employed to increase enzyme production, including the use of molecular tools, strain engineering, and in silico optimization. The review article highlights the critical role that rational design plays in achieving successful heterologous expression and underscores the challenges of large-scale production of L-ASNase, such as inadequate protein folding and the metabolic burden on host cells. Improved gene expression is shown to be achievable through the optimization of codon usage, synthetic promoters, transcription and translation regulation, and host strain improvement, among others. Additionally, this review provides a deep understanding of the enzymatic properties of L-ASNase and how this knowledge has been employed to enhance its properties and production. Finally, future trends in L-ASNase production, including the integration of CRISPR and machine learning tools are discussed. This work serves as a valuable resource for researchers looking to design effective heterologous expression systems for L-ASNase production as well as for enzymes production in general.

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Section: Type and Characteristics Of L-asnase In Organismsmentioning

confidence: 99%

Section: Improvement In Systems For L-asnase Heterologous Expressionmentioning

confidence: 99%

Current state of molecular and metabolic strategies for the improvement of L-asparaginase expression in heterologous systems

et al. 2023

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“…The same doubt was expressed about the type I enzyme of archaea from Pyrococcus horikoshi , which acts as a dimer rather than a tetramer [ 33 ]. L-ASNases of some extremophile bacterial organisms [ 61 ], for example, Thermus thermophilus [ 62 ], Thermococus sibiricus [ 63 ] or Melioribacter roseus [ 64 ], can act as hexamers (trimers of dimers), but there is no structural evidence to support this hypothesis. It is believed that the dimer of L-ASNase II, which has two active centers, is not able to cleave L-asparagine.…”

Section: Structures Of L-asnases and The Mechanism Of Actionmentioning

confidence: 99%

Molecular Analysis of L-Asparaginases for Clarification of the Mechanism of Action and Optimization of Pharmacological Functions

et al. 2022

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L-asparaginases (EC 3.5.1.1) are a family of enzymes that catalyze the hydrolysis of L-asparagine to L-aspartic acid and ammonia. These proteins with different biochemical, physicochemical and pharmacological properties are found in many organisms, including bacteria, fungi, algae, plants and mammals. To date, asparaginases from E. coli and Dickeya dadantii (formerly known as Erwinia chrysanthemi) are widely used in hematology for the treatment of lymphoblastic leukemias. However, their medical use is limited by side effects associated with the ability of these enzymes to hydrolyze L-glutamine, as well as the development of immune reactions. To solve these issues, gene-editing methods to introduce amino-acid substitutions of the enzyme are implemented. In this review, we focused on molecular analysis of the mechanism of enzyme action and to optimize the antitumor activity.

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“…After optimizing induction and purification conditions, the L-ASNase reached a specific activity of 312.8 U/mg wich means in 1.5 times control activity. In a work by Dumina et al (2021), the activity of new, uncharacterized extremophilic L-ASNases of the psychrophilic fungus Sclerotinia borealis , the thermoacidophilic crenarchae-onAcidilobus saccharovorans and the thermophilic bacteriumMelioribacter roseu were studied. These were expressed inE.…”

Section: Improvement In Systems For L-asnase Heterologous Expressionmentioning

confidence: 99%

Current state of molecular and metabolic strategies for the improvement of L-asparaginase expression in heterologous systems

Lefin

Miranda

Beltrán

et al. 2022

Preprint

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L-asparaginase is one of the world’s most in-demand industrial enzymes, mainly due to its therapeutic properties and its use in the food industry. Over the years studies have been conducted to improve its specific activity and reduce side effects, driving new discoveries that promise safer, more efficient, and cheaper drugs for consumers. However, heterologous protein modifications or expression continue to be a problem during production. Therefore, addressing bottlenecks when attempting to express modified and/or heterologous proteins using the rational design of heterologous expression systems with modified hosts could be a promising strategy to improve L-asparaginase production. Problems such as inadequate protein folding, metabolic load on host cells, codon usage bias, repetitive sequences affecting translation, heavy molecular weight and/or multi-membrane domains formation; need great attention during the development of “bio-better” proteins. In this article, we address the current molecular and metabolic strategies that have been developed to improve the heterologous expression of L-asparaginase.

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Comparison of Enzymatic Activity of Novel Recombinant L-asparaginases of Extremophiles

Cited by 9 publications

References 39 publications

Current state of molecular and metabolic strategies for the improvement of L-asparaginase expression in heterologous systems

Current state of molecular and metabolic strategies for the improvement of L-asparaginase expression in heterologous systems

Molecular Analysis of L-Asparaginases for Clarification of the Mechanism of Action and Optimization of Pharmacological Functions

Current state of molecular and metabolic strategies for the improvement of L-asparaginase expression in heterologous systems

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