Structural aspects of L-asparaginases, their friends and relations.

Michalska, K.; Jaskólski, M.

doi:10.18388/abp.2006_3291

Cited by 116 publications

(80 citation statements)

References 71 publications

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“…The 16S rRNA sequencing was found to be a phylogenetic tool also useful for bacterial detection and identification, it can better identify poorly described strains (Matsumoto and Sugano Fortunately, our test isolate that used in the present study could produce l-asparaginase extracellularly (Type II l-asparaginase) at a level 470% more than its intracellular level. Bacterial l-asparaginases are defined by their intra or extra cellular localization (Michalska and Jaskolski 2006). Type I (cytosolic) binds to l-asparagine with low affinity, in contrast the extracellular type II (periplasmic) has a strong binding capacity and it has a characteristic oligomeric form.…”

Section: Discussionmentioning

confidence: 99%

Production, characterization and bioinformatics analysis of l-asparaginase from a new Stenotrophomonas maltophilia EMCC2297 soil isolate

et al. 2020

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An exhaustive screening program was applied for scoring a promising l-asparaginase producing-isolate. The recovered isolate was identified biochemically and molecularly and its l-asparaginase productivity was optimized experimentally and by Response Surface Methodology. The produced enzyme was characterized experimentally for its catalytic properties and by bioinformatics analysis for its immunogenicity. The promising l-asparaginase producing-isolate was selected from 722 recovered isolates and identified as Stenotrophomonas maltophilia and deposited at Microbiological Resources Centre (Cairo Mircen) under the code EMCC2297. This isolate produces both intracellular (type I) and extracellular (type II) l-asparaginases with about 4.7 fold higher extracellular l-asparaginase productivity. Bioinformatics analysis revealed clustering of Stenotrophomonas maltophilia l-asparaginase with those of Pseudomonas species and considerable closeness to the two commercially available l-asparaginases of E. coli and Erwinia chrysanthemi. Fourteen antigenic regions are predicted for Stenotrophomonas maltophilia l-asparaginase versus 16 and 18 antigenic regions for the Erwinia chrysanthemi and E. coli l-asparaginases. Type II l-asparaginase productivity of the test isolate reached 4.7 IU/ml/h and exhibited maximum activity with no metal ion requirement at 37 °C, pH 8.6, 40 mM asparagine concentration and could tolerate NaCl concentration up to 500 mM and retain residual activity of 55% at 70 °C after half an hour treatment period. Application both of random mutation by gamma irradiation and Response Surface Methodology that determined 38.11 °C, 6.89 pH, 19.85 h and 179.15 rpm as optimum process parameters could improve the isolate l-asparaginase productivity. Maximum production of about 8 IU/ml/h was obtained with 0.4% dextrose, 0.1% yeast extract and 10 mM magnesium sulphate. In conclusion l-asparaginase of the recovered Stenotrophomonas maltophilia EMCC2297 isolate has characters enabling it to be used for medical therapeutic application.

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Section: Discussionmentioning

confidence: 99%

Production, characterization and bioinformatics analysis of l-asparaginase from a new Stenotrophomonas maltophilia EMCC2297 soil isolate

et al. 2020

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“…Another functional group in the active site that can be ionized is the aamino group of the Thr-196 nucleophile. This group needs to be unprotonated in Ntn hydrolases in order to activate the side-chain hydroxyl for nucleophilic attack on the amide group of the substrate [3]. The value of pK 1 is most likely to reflect the ionization of this group.…”

Section: Discussionmentioning

confidence: 99%

“…L-Asparaginase is the enzyme responsible for this hydrolytic reaction. L-Asparaginases are present in most organisms, and have been classified into two major types, the bacterial-(also called type I and II) and plant-type (or type III) asparaginases [3]. The bacterial type II enzymes, with high L-asparagine affinity, are useful for treating acute lymphoblastic leukemia [4].…”

Section: Introductionmentioning

confidence: 99%

Structural basis of potassium activation in plant asparaginases

et al. 2018

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l-asparaginases (EC 3.5.1.1) play an important role in nitrogen mobilization in plants. Here, we investigated the biochemical and biophysical properties of potassium-dependent (PvAspG1) and potassium-independent (PvAspG-T2) l-asparaginases from Phaseolus vulgaris. Our previous studies revealed that PvAspG1 requires potassium for catalytic activation and its crystal structure suggested that Ser-118 in the activation loop plays a critical role in coordinating the metal cation. This amino acid residue is replaced by isoleucine in PvAspG-T2. Reciprocal mutants of the enzymes were produced and the effect of the amino acid substitution on the kinetic parameters, allosteric effector binding, secondary structure conformation, and pH profile were studied. Introduction of the serine residue conferred potassium activation in PvAspG-T2. Conversely, the PvAspG1-S118I mutant could no longer be activated by potassium. PvAspG1 and the PvAspG-T2-I117S mutant had a similar half-maximal effective concentration (EC ) value for potassium activation, between 0.1 and 0.3 mm. Potassium binding elicited a similar conformational change in PvAspG1 and PvAspG-T2-I117S, as studied by circular dichroism. However, no change in conformation was observed for PvAspG-T2 and PvAspG1-S118I. Analysis of kinetic parameters in function of pH indicated that potassium activation mediated by Ser-118 influences the ionization of specific functional groups in the enzyme-substrate complex. Together, the results indicate that Ser-118 of PvAspG1 is essential and sufficient for potassium activation in plant l-asparaginases. ENZYME: l-Asparaginase (EC 3.5.1.1).

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“…In E. coli , two evolutionarily related l ‐asparaginases are present: the periplasmic, highly active enzyme ( K M = 10–12 μ m ) with potent cytotoxic properties, and its less active but allosteric cytosolic homolog ( K 0.5 = 1.0 m m ) lacking any antileukemic properties . The two enzymes have been classified as bacterial‐type l ‐asparaginases type I (the cytoplasmic protein, EcAI) and type II (the periplasmic protein, EcAII) to differentiate them from each other as well as from another group of evolutionarily unrelated l ‐asparaginases, sometimes called plant‐type l ‐asparaginases . The l ‐asparaginase from E. chrysanthemi and EcAII were soon introduced into clinical practice as very effective agents for the treatment of childhood acute lymphoblastic leukemia (ALL) and became standard components of protocols in ALL therapy .…”

Section: Introductionmentioning

confidence: 99%

Crystal structure of active site mutant of antileukemic l‐asparaginase reveals conserved zinc‐binding site

et al. 2014

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L-aspartate and ammonia. EcAII is an important drug in the treatment of childhood acute lymphoblastic leukemia, the most common malignancy in children. Leukemic blast cells lack the ability to synthesize L-asparagine and rely on other sources of L-asparagine for protein synthesis. EcAII injections deplete extracellular levels of L-asparagine, disrupting protein synthesis and inducing apoptosis in the malignant cells. The detailed mechanism of L-asparaginase catalytic action, the molecular mechanisms of its anticancer activity and the side effects associated with the treatment, including resistance to therapy, are not fully understood despite over 40 years of research. Here, we present X-ray structures of EcAII with an active site mutation, D90E, in three crystal forms. The region of the mutation is well ordered, allowing precise functional analysis of the consequences of the replacement of Asp90. In all three structures, the mutant protein exhibits an open conformation of the active site. In one of the structures, a zinc cation has been detected. The zinc cation is coordinated in a region of the protein that is implicated in the immunological response to EcAII treatment. A combined sequence-structure analysis of bacterial-type L-asparaginases reveals that the metal coordination may play a role in the response to asparaginase treatment. The observation of a zinc-binding site in antileukemic L-asparaginases provides new insight, with consequences for acute lymphoblastic leukemia therapy. DatabasesThe atomic coordinates of the monoclinic, orthorhombic and trigonal forms of the D90E mutant of Escherichia coli type II asparaginase have been deposited with the RCSB PDB with accession codes 1JAZ, 1JJA and 1IHD, respectively. Enzymes EC 3.5

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Structural aspects of L-asparaginases, their friends and relations.

Cited by 116 publications

References 71 publications

Production, characterization and bioinformatics analysis of l-asparaginase from a new Stenotrophomonas maltophilia EMCC2297 soil isolate

Production, characterization and bioinformatics analysis of l-asparaginase from a new Stenotrophomonas maltophilia EMCC2297 soil isolate

Structural basis of potassium activation in plant asparaginases

Crystal structure of active site mutant of antileukemic l‐asparaginase reveals conserved zinc‐binding site

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