[4] The N-terminal region consists of two layers of six-stranded antiparallel β-sheets between which is the active site responsible for the hydrolysis of glutamine.

[4] These two active sites are connected by a tunnel lined primarily with backbone atoms and hydrophobic, nonpolar amino acid residues.

[4] Structural characterization of asparagine synthetase from mammalian sources have been difficult due to the low abundance and instability of the enzyme during purification procedures.

[5] Thus, after being released in, and channeled from, the glutaminase site, the ammonia molecule attacks the bound βAspAMP 1 to give asparagine and AMP via a tetrahedral intermediate.

[5] Other experiments demonstrated that quiescent rat thyroid cells entering S phase as a result of thyroid-stimulating hormone treatment was matched with a concurrent increase in asparagine synthetase mRNA content.

[3] For example, in mouse models, 24 hours after exposure to L-asparaginase, tumors resistant to the depletion responded with 5- to 19-fold increases in asparagine synthetase expression.

[14] It has been further demonstrated in mouse model systems that repeated subculturing of L-asparaginase sensitive tumor cells in sublethal concentrations of L-asparaginase could eventually make them resistant, a potential worry of lower dose chemotherapy treatments effectively encouraging resistant cell development.

[15] A correlation between L-asparaginase efficacy and asparagine synthetase protein levels in a number of human ovarian cell lines has been observed.

[2][18] These circulating tumor cells were also found to have an increased capacity for colony formation in soft agar assays under hypoxic conditions and grew faster when reimplanted as xenografts.