Three-dimensional structures of<scp>L</scp>-asparaginase from<i>Erwinia carotovora</i>complexed with aspartate and glutamate

Кравченко, О. В.; Kislitsin, Y.A.; Попов, А. Н.; Nikonov, S.V.; Куранова, И. П.

doi:10.1107/s0907444907065766

Cited by 22 publications

(16 citation statements)

References 28 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…This model, however, proved useful in molecular replacement trials that led to the crystal structure determination of EcAII at 2.3 Å resolution in 1993 [34]. Subsequently, crystal structures of a number of LASNases were determined, including ErA [35], Wolinella succinogenes L-ASNase (WsA) [36], Erwinia carotovora (EwA) [37,38] and related asparaginase-glutaminases from Acinetobacter glutaminasificans (AgA) [23] and Pseudomonas fluorescence (PgA) [22] as shown in Fig. (3).…”

Section: The Three Dimensional Structure Of L-asnasementioning

confidence: 99%

Structure-Function Relationships and Clinical Applications of L-Asparaginases

Labrou¹,

Papageorgiou²,

Avramis³

2010

CMC

View full text Add to dashboard Cite

L-asparaginase (L-ASNase, EC 3.5.1.1) catalyzes the hydrolysis of the non-essential amino acid L-Asn to LAsp and ammonia and is widely used for the treatment of haematopoetic diseases such as acute lymphoblastic leukaemia (ALL) and lymphomas. Therapeutic forms of L-ASNase come from different biological sources (primarily E. coli and Erwinia chrysanthemi). It is well established that the various preparations have different biochemical pharmacology properties, and different tendency to induce side-effects. This is due to different structural, physicochemical and kinetic properties of L-ASNases from the various biological sources. Understanding these properties of various L-ASNases would allow a better decipherment of their catalytic and therapeutic features, thus enabling more accurate predictions of the behaviour of these enzymes under a variety of therapeutic conditions. In addition, detailed understanding of the catalytic mechanism of L-ASNases might permit the design of new forms of L-ASNases with optimal biochemical properties for clinical applications. In this paper we review the available biochemical and pharmacokinetic information of the therapeutic forms of bacterial L-ASNases, and focus on a detailed description of structure, function and clinical applications of these enzymes.

show abstract

Section: The Three Dimensional Structure Of L-asnasementioning

confidence: 99%

Structure-Function Relationships and Clinical Applications of L-Asparaginases

Labrou¹,

Papageorgiou²,

Avramis³

2010

CMC

View full text Add to dashboard Cite

show abstract

“…The wild-type enzyme is also effective against cells with low ASNS expression [17]. A better understanding of substrate specificity may provide a path to optimizing asparaginase therapy [18][19][20][21][22] by engineering analogs with tailored ratios of asparaginase:glutaminase activity. .…”

Section: Introductionmentioning

confidence: 99%

Catalytic Role of the Substrate Defines Specificity of Therapeutic l-Asparaginase

Anishkin

Vanegas

Rogers

et al. 2015

Journal of Molecular Biology

View full text Add to dashboard Cite

Type II bacterial L-asparaginases (L-ASP) have played an important therapeutic role in cancer treatment for over four decades, yet their exact reaction mechanism remains elusive. L-ASP from Escherichia coli deamidates asparagine (Asn) and glutamine, with an ~10(4) higher specificity (kcat/Km) for asparagine despite only one methylene difference in length. Through a sensitive kinetic approach, we quantify competition among the substrates and interpret its clinical role. To understand specificity, we use molecular simulations to characterize enzyme interactions with substrates and a product (aspartate). We present evidence that the aspartate product in the crystal structure of L-ASP exists in an unusual α-COOH protonation state. Consequently, the set of enzyme-product interactions found in the crystal structure, which guided prior mechanistic interpretations, differs from those observed in dynamic simulations of the enzyme with the substrates. Finally, we probe the initial nucleophilic attack with ab initio simulations. The unusual protonation state reappears, suggesting that crystal structures (wild type and a T89V mutant) represent intermediate steps rather than initial binding. Also, a proton transfers spontaneously to Asn, advancing a new hypothesis that the substrate's α-carboxyl serves as a proton acceptor and activates one of the catalytic threonines during L-ASP's nucleophilic attack on the amide carbon. That hypothesis explains for the first time why proximity of the substrate α-COO(-) group to the carboxamide is absolutely required for catalysis. The substrate's catalytic role is likely the determining factor in enzyme specificity as it constrains the allowed distance between the backbone carboxyl and the amide carbon of any L-ASP substrate.

show abstract

“…The L-asparaginase template for homology modelling was identified on the basis of criteria such as maximum score, smaller the e-value, >30% identity with other L-asparaginases (from Escherichia coli 11 , Wolinella succinogenes 12 , Erwinia chrysanthemi 13 , Erwinia carotovora 14 ) reported in the literature. It is also reported in the literature that the L-asparaginase from above bacterial strains shows high similarity with respect to tertiary and quaternary structures.…”

Section: Results and Discussion: Homology Modelling And In-silico Mutmentioning

confidence: 99%

“…3b to both the enzymes of L-asparaginases (wild and mutated version) were performed by using auto dock tool (ADT) 4.2 (http://autodock.scripps.edu/ resources/adt). The ADT graphics interface was employed for manual preparation of the protein by adding polar hydrogens and merging non-polar hydrogens 7 . In the case of ligand, both the ligands were sketched in Tripo's Sybyl6.7 and Gasteiger-Huckel charges were added to minimise the molecules and non-polar hydrogens were merged to give stability.…”

Section: In-silico Mutagenesismentioning

confidence: 99%

Untitled

2018

IJPSR

View full text Add to dashboard Cite

L-asparaginase has been accepted clinically as an anti-tumour agent for the effective treatment of acute lymphoblastic leukaemia and lymphosarcoma. This enzyme also possesses L-glutaminase activity and causes immunological problems. Hence, efforts have been made to develop mutants with lower or no glutaminase activity. In the present study, a homology model of L-asparaginase obtained from Pectobacterium carotovorum was docked and compared with its mutants for activities, analysed for molecular dynamics and structural stability. A total of five in-silico mutants were developed using single amino acid mutagenesis and evaluated for ligand binding for both L-asparagine and L-glutamine. One of the mutants, Y306L, showed -5.89 and -5.04 while the wild L-asparaginase revealed -5.50 and -5.12 binding energies for L-asparagine and L-glutamine, respectively. Further, substrate-enzyme interaction analysis indicated that the wild L-asparaginase showed five interactions for L-asparagine as well as for L-glutamine, whereas, the mutant, Y306L depicted eight and three interactions for L-asparagine and L-glutamine. Molecular dynamics analysis denoted that mutant protein is more stable in RMSD, RMSF and radius of gyration to that of wild-type protein. Higher binding affinity was noticed with Lasparagine and lower with L-glutamine along with higher interactions with Lasparagine and lower with L-glutamine in mutant Y306L, compared to wild Lasparaginase. This suggests that this could be a possible potential candidate for the treatment of Acute Lymphoblastic Leukaemia (ALL) with less induced side effects.

show abstract

Three-dimensional structures ofL-asparaginase fromErwinia carotovoracomplexed with aspartate and glutamate

Cited by 22 publications

References 28 publications

Structure-Function Relationships and Clinical Applications of L-Asparaginases

Structure-Function Relationships and Clinical Applications of L-Asparaginases

Catalytic Role of the Substrate Defines Specificity of Therapeutic l-Asparaginase

Untitled

Contact Info

Product

Resources

About