Structural and biophysical aspects of <scp>L</scp>-asparaginases: a growing family with amazing diversity

Loch, J.I.; Jaskólski, Mariusz

doi:10.1107/s2052252521006011

Cited by 24 publications

(14 citation statements)

References 120 publications

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“…One report suggests that the total ASNase activity in a lysate of cells from the wild strain of E. coli corresponds to very low micromolar concentrations of ASNases [20]. Since E. coli encodes several enzymes capable of l ‐Asn hydrolysis [7], the concentration of EcAI alone could well be in the picomolar range. EcAI is highly homologous to YpAI and structurally nearly indistinguishable.…”

Section: Discussionmentioning

confidence: 99%

“…ASNases ( l ‐asparaginases; E.C.3.5.1.1 or l ‐glutaminase‐asparaginases; E.C.3.5.1.38) catalyse the hydrolysis of l ‐Asn to l ‐Asp, with varied additional ability to utilize other substrates, such as l ‐Gln, d ‐Asn, d ‐Gln, succinamic acid, etc. Various criteria have been used to classify ASNases [7,8]. In this report, we categorize these enzymes based strictly on their structural properties [1].…”

Section: Introductionmentioning

confidence: 99%

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The dimeric form of bacterial l‐asparaginase YpAI is fully active

et al. 2022

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l‐asparaginases from mesophilic bacteria (ASNases), including two enzymes very successfully used in the treatment of leukaemia, have been consistently described as homotetramers. On the contrary, structural studies show that homodimers of these enzymes should be sufficient to carry out the catalytic reaction. In this report, we investigated whether the type I Yersinia pestis asparaginase (YpAI) is active in a dimeric form or whether the tetrameric quaternary structure is critical for its activity. Using multiple biophysical techniques that investigate enzymatic properties and quaternary structure at either high or low protein concentration, we found that dimeric YpAI is fully active, suggesting that the tetrameric form of this subfamily of enzymes does not bear significant enzymatic relevance. In this process, we extensively characterized YpAI, showing that it is a cooperative enzyme, although the mechanism of allostery is still not definitely established. We showed that, like most type I ASNases, the substrate affinity of YpAI is low and this enzyme is very similar in terms of both the structure and enzymatic properties to homologous type I ASNase from Escherichia coli (EcAI). We extended these studies to more medically relevant type II ASNases, used as anti‐leukaemia drugs. We confirmed that type II ASNases are not allosteric, and that they might also be functional in a dimeric form. However, the determination of the accurate tetramer⇆dimer dissociation constants of these enzymes that most likely lie in the picomolar range is not possible with currently available biophysical techniques.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

The dimeric form of bacterial l‐asparaginase YpAI is fully active

et al. 2022

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“…Recent data suggest that EcAII enables anaerobic respiration and is part of a metabolic pathway leading to the production of fumarate as a terminal electron acceptor. is localized in the periplasm to be able to utilize exogenous (extracellular) L-Asn to produce fumarate, as under anaerobic conditions transport of L-Asn from the cytoplasm to the periplasm is abolished [21,40].…”

Section: Discussionmentioning

confidence: 99%

“…The properties of R. etli L-ASNases have yet to be elucidated. However, preliminary kinetic data indicate that these enzymes display submillimolar K M and no glutaminase activity, and thus can be considered as potential antileukemic agents [21].…”

Section: Introductionmentioning

confidence: 99%

Highly Active Thermophilic L-Asparaginase from Melioribacter roseus Represents a Novel Large Group of Type II Bacterial L-Asparaginases from Chlorobi-Ignavibacteriae-Bacteroidetes Clade

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Zhgun

Pokrovskaya

et al. 2021

IJMS

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L-asparaginase (L-ASNase) is a biotechnologically relevant enzyme for the pharmaceutical, biosensor and food industries. Efforts to discover new promising L-ASNases for different fields of biotechnology have turned this group of enzymes into a growing family with amazing diversity. Here, we report that thermophile Melioribacter roseus from Ignavibacteriae of the Bacteroidetes/Chlorobi group possesses two L-ASNases—bacterial type II (MrAII) and plant-type (MrAIII). The current study is focused on a novel L-ASNase MrAII that was expressed in Escherichia coli, purified and characterized. The enzyme is optimally active at 70 °C and pH 9.3, with a high L-asparaginase activity of 1530 U/mg and L-glutaminase activity ~19% of the activity compared with L-asparagine. The kinetic parameters KM and Vmax for the enzyme were 1.4 mM and 5573 µM/min, respectively. The change in MrAII activity was not significant in the presence of 10 mM Ni2+, Mg2+ or EDTA, but increased with the addition of Cu2+ and Ca2+ by 56% and 77%, respectively, and was completely inhibited by Zn2+, Fe3+ or urea solutions 2–8 M. MrAII displays differential cytotoxic activity: cancer cell lines K562, Jurkat, LnCap, and SCOV-3 were more sensitive to MrAII treatment, compared with normal cells. MrAII represents the first described enzyme of a large group of uncharacterized counterparts from the Chlorobi-Ignavibacteriae-Bacteroidetes clade.

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“…The practical application of L-ASNases is currently directly related to their enzymatic properties. The structural, biophysical and biochemical properties of type I and II L-ASNases have been described in experimental works [ 11 , 27 , 28 , 29 , 30 ], and detailed information about the catalytic mechanism of bacterial L-ASNases is presented in detail in the reviews by Lubkowski et al [ 31 ] and Loch [ 32 ].…”

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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Structural and biophysical aspects of L-asparaginases: a growing family with amazing diversity

Cited by 24 publications

References 120 publications

The dimeric form of bacterial l‐asparaginase YpAI is fully active

The dimeric form of bacterial l‐asparaginase YpAI is fully active

Highly Active Thermophilic L-Asparaginase from Melioribacter roseus Represents a Novel Large Group of Type II Bacterial L-Asparaginases from Chlorobi-Ignavibacteriae-Bacteroidetes Clade

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

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