Study on the function of endogenous opioid and substance P at inflammation and pain

我们合成了Spinorphin类似物，并测定了它们对脑啡肽降解酶中的二肽基肽酶III（DPPIII）的抑制活性。VVYPW是Spinorphin的一种N端和C端截短形式，具有更强的抑制活性。这一结果表明VVYPW具有更有效的表达DPPIII抑制活性的结构。我们试图将伤害性感觉纤维分为三种类型，并比较Spinorphin和吗啡诱导的镇痛作用模式。因此，spinorphin完全阻断2-methyl-thioadenosine引起的反应，而吗啡则无此作用。另一方面，吗啡诱导的缓激肽反应的阻滞被百日咳毒素处理减弱，而Spinorphin的则没有。在人P物质受体的纯化方面，我们构建了N端含6xHis表位的重组P物质受体。6xHis标签不干扰受体结合配体的能力，6XHis-SPR可以在Ni-NTA柱上纯化。该纯化过程构成了获得大量均质SPR用于结构研究的重要步骤。

We synthesized spinorphin analogous and assayed their inhibitoriy activity toward dipeptidyl peptidase III (DPPIII) among enkephalin degrading enzymes. VVYPW, an N-terminal and C-terminal truncated form of spinorphin, exhibited more potent inhibitory activity.This results indicated that VVYPW had amore effective structure of expression of inhibitory activity toward DPPIII.We attempted to characterize nociceptive sensory fibers into three types, and examined the mode of action of spinorphin-induced analgesia, in comparison to morphine-induced one.So, spinorphin completely blocked 2-metyl-thioadenosine induced responsed, but morphine did not. On the other hand, morphine-induced blockade of bradykinin responses was attenuated by pertussis toxin treatment, while the spinorphin's ones was not. Thus it is suggested that spinorphin has wide spectrum of analgesia which covers the blockade of nociception insensitive to morphine.Regarding to purification of human substance P receptor, we have developed a recombinant Substance P receptor with 6xHis epitope at its N-terminus. The 6xHis tag does not interfere with receptor's ability to bind ligands the 6XHis-SPR can be purified over the Ni-NTA column. This purification procedure constitutes an important step toward obtaining large quantities of homogeneous SPR for structural studies.