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Crystal structure of active site mutant of antileukemic l ‐asparaginase reveals conserved zinc‐binding site
Author(s) -
Borek Dominika,
Kozak Maciej,
Pei Jimin,
Jaskolski Mariusz
Publication year - 2014
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.12906
Subject(s) - asparagine , active site , asparaginase , periplasmic space , chemistry , mutant , mutation , biochemistry , biology , cancer research , enzyme , escherichia coli , leukemia , genetics , gene , lymphoblastic leukemia
The periplasmic enzyme l ‐asparaginase type II from Escherichia coli (Ec AII ) converts l ‐asparagine to l ‐aspartate and ammonia. Ec AII 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. Ec AII 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 Ec AII 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 Ec AII 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. Databases The 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 3.5.1.1 , l ‐asparagine amidohydrolase, l ‐asparaginase; EC 3.5.1.38 , l ‐glutamine ( l ‐asparagine) amidohydrolase, glutaminase‐asparaginase; EC 6.3.5.6 , l ‐aspartyl‐tRNA Asn : l ‐glutamine amido‐ligase (ADP‐forming), asparaginyl‐tRNA synthase (glutamine‐hydrolysing).