Jain Rohit, Khan Nazimuddin, Menzel Andreas, Rajkovic Ivan, Konrad Manfred, Techert Simone
Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, 37077, Göttingen, Germany.
Paul Scherrer Institute, 5232, Villingen, Switzerland.
Eur Biophys J. 2016 Jan;45(1):81-9. doi: 10.1007/s00249-015-1079-9. Epub 2015 Oct 7.
Bio-catalysis is the outcome of a subtle interplay between internal motions in enzymes and chemical kinetics. Small-angle X-ray scattering (SAXS) investigation of an enzyme's internal motions during catalysis offers an integral view of the protein's structural plasticity, dynamics, and function, which is useful for understanding allosteric effects and developing novel medicines. Guanylate kinase (GMPK) is an essential enzyme involved in the guanine nucleotide metabolism of unicellular and multicellular organisms. It is also required for the intracellular activation of numerous antiviral and anticancer purine nucleoside analog prodrugs. Catalytically active recombinant human GMPK (hGMPK) was purified for the first time and changes in the size and shape of open/closed hGMPK were tracked by SAXS. The binding of substrates (GMP + AMPPNP or Ap5G or GMP + ADP) resulted in the compaction of size and shape of hGMPK. The structural changes between open and completely closed hGMPK conformation were confirmed by observing differences in the hGMPK secondary structures with circular dichroism spectroscopy.
生物催化是酶内部运动与化学动力学之间微妙相互作用的结果。小角X射线散射（SAXS）对酶催化过程中内部运动的研究提供了蛋白质结构可塑性、动力学和功能的整体视图，这对于理解变构效应和开发新药很有用。鸟苷酸激酶（GMPK）是单细胞和多细胞生物鸟嘌呤核苷酸代谢中必不可少的一种酶。它也是许多抗病毒和抗癌嘌呤核苷类似物前药细胞内活化所必需的。首次纯化了具有催化活性的重组人GMPK（hGMPK），并通过SAXS追踪开放/闭合hGMPK的大小和形状变化。底物（GMP + AMPPNP或Ap5G或GMP + ADP）的结合导致hGMPK的大小和形状压缩。通过圆二色光谱观察hGMPK二级结构的差异，证实了开放和完全闭合hGMPK构象之间的结构变化。
