Division of Medicine, University College London, Gower Street, London WC1E 6BT, England.
School of Biological Sciences, University of the Punjab, Quaid-e-Azam Campus, Lahore 54590, Pakistan.
Acta Crystallogr D Struct Biol. 2017 Nov 1;73(Pt 11):889-895. doi: 10.1107/S2059798317014711. Epub 2017 Oct 20.
L-Asparaginases catalyse the hydrolysis of asparagine to aspartic acid and ammonia. In addition, L-asparaginase is involved in the biosynthesis of amino acids such as lysine, methionine and threonine. These enzymes have been used as chemotherapeutic agents for the treatment of acute lymphoblastic leukaemia and other haematopoietic malignancies since the tumour cells cannot synthesize sufficient L-asparagine and are thus killed by deprivation of this amino acid. L-Asparaginases are also used in the food industry and have potential in the development of biosensors, for example for asparagine levels in leukaemia. The thermostable type I L-asparaginase from Thermococcus kodakarensis (TkA) is composed of 328 amino acids and forms homodimers in solution, with the highest catalytic activity being observed at pH 9.5 and 85°C. It has a K value of 5.5 mM for L-asparagine, with no glutaminase activity being observed. The crystal structure of TkA has been determined at 2.18 Å resolution, confirming the presence of two α/β domains connected by a short linker region. The N-terminal domain contains a highly flexible β-hairpin which adopts open' and closed' conformations in different subunits of the solved TkA structure. In previously solved L-asparaginase structures this β-hairpin was only visible when in the `closed' conformation, whilst it is characterized with good electron density in all of the subunits of the TkA structure. A phosphate anion resides at the active site, which is formed by residues from both of the neighbouring monomers in the dimer. The high thermostability of TkA is attributed to the high arginine and salt-bridge content when compared with related mesophilic enzymes.
L-天冬酰胺酶催化天冬酰胺水解为天冬氨酸和氨。此外，L-天冬酰胺酶还参与赖氨酸、蛋氨酸和苏氨酸等氨基酸的生物合成。由于肿瘤细胞不能合成足够的 L-天冬酰胺，因此会因缺乏这种氨基酸而死亡，这些酶已被用作治疗急性淋巴细胞白血病和其他血液恶性肿瘤的化疗药物。L-天冬酰胺酶也用于食品工业，并有潜力开发用于生物传感器，例如用于白血病中天冬酰胺水平的传感器。来自 Thermococcus kodakarensis (TkA) 的耐热型 I 型 L-天冬酰胺酶由 328 个氨基酸组成，在溶液中形成同源二聚体，在 pH 9.5 和 85°C 时观察到最高的催化活性。它对 L-天冬酰胺的 K 值为 5.5 mM，没有观察到谷氨酰胺酶活性。TkA 的晶体结构已在 2.18 Å 分辨率下确定，证实存在两个通过短连接区连接的 α/β 结构域。N 端结构域包含一个高度灵活的β发夹，在已解决的 TkA 结构的不同亚基中采用“打开”和“关闭”构象。在以前解决的 L-天冬酰胺酶结构中，只有当处于“关闭”构象时，才能看到这个β发夹，而在 TkA 结构的所有亚基中，它的特征是具有良好的电子密度。一个磷酸阴离子位于活性位点，由二聚体中相邻两个单体的残基形成。与相关的嗜温酶相比，TkA 的高耐热性归因于高精氨酸和盐桥含量。
