Asparagine synthetase: Function, structure, and role in disease

Abstract

Asparagine synthetase (ASNS) converts aspartate and glutamine to asparagine and glutamate in an ATP-dependent reaction. ASNS is present in most, if not all, mammalian organs, but varies widely in basal expression. Human ASNS activity is highly responsive to cellular stress, primarily by increased transcription from a single gene located on chromosome 7. Elevated ASNS protein expression is associated with resistance to asparaginase therapy in childhood acute lymphoblastic leukemia. There is evidence that ASNS expression levels may also be inversely correlated with asparaginase efficacy in certain solid tumors as well. Children with mutations in the ASNS gene exhibit developmental delays, intellectual disability, microcephaly, intractable seizures, and progressive brain atrophy. Thus far, 15 unique mutations in the ASNS gene have been clinically associated with asparagine synthetase deficiency (ASD). Molecular modeling using the Escherichia coli ASNS-B structure has revealed that most of the reported ASD substitutions are located near catalytic sites or within highly conserved regions of the protein. For some ASD patients, fibroblast cell culture studies have eliminated protein and mRNA synthesis or stability as the basis for decreased proliferation.

Keywords: acute lymphoblastic leukemia; amino acid; amino acid metabolism; asparaginase resistance; brain development; brain metabolism; genetic disease; inborn error of metabolism; neurological disease; protein structure.

Figure 1.

Mechanism and structural features of human asparagine synthetase. A, the reaction begins when the aspartate carboxyl is activated by an ATP-dependent process, forming a β-aspartyl-AMP intermediate. Glutamine deamidation releases ammonia, which performs a nucleophilic attack on the aspartyl intermediate to produce asparagine. Glutamate is the second product of the overall reaction. B, sequence of human ASNS isoform 1 with residues colored light and dark gray for the N- and C-terminal domains, respectively. α-Helical and β-sheet secondary structures are shown above the sequence. Residues associated with glutamine and ATP binding are colored purple and orange, respectively. Clinically identified ASD mutations are colored according to Fig. 3. C, the N- and C-terminal domains are represented within the surface structure and colored light and dark gray, respectively. The substrates glutamine (purple) and ATP (orange) are indicated by an arrow and shown as sticks. Insets show the substrate-binding pockets. Hydrogen bonds are shown as black dashes with distances labeled in angstroms.

Figure 2.

Regulation of ASNS expression. Asparagine depletion activates the AAR, whereas endoplasmic reticulum stress (ER Stress) activates the UPR. Each stress condition increases the activity of an eIF2 kinase. Phosphorylation of eIF2 slows global protein synthesis, but paradoxically increases translation of a subset of mRNAs, including that for the transcription factor ATF4. Binding of ATF4 to an enhancer element within the promoter of the ASNS gene induces expression of the enzyme.

Figure 3.

ASD associated mutations in the human ASNS enzyme. Mutations are represented as sticks within the predicted ASNS structure. Top panel, A6E (red), L145S (salmon), T337I (light green), R340H (dark green), A380S (light blue), and Y398C (dark blue). Bottom panel (180° rotation around y axis), R49Q (raspberry), L247W (light orange), G289A (yellow), F362V (teal), S480F (light pink), and V489D (dark pink). Interacting residues are also shown as sticks, and hydrogen bonds are represented as black dashes.