Gilbert J A, Hanson P D, McCormick D J, Richelson E
Department of Psychiatry, Mayo Clinic and Foundation, Rochester, Minnesota 55905.
J Neurochem. 1987 Dec;49(6):1845-52. doi: 10.1111/j.1471-4159.1987.tb02446.x.
Murine neuroblastoma clone N1E-115, which possesses receptors for neurotensin mediating the formation of intracellular cyclic GMP and the stimulation of inositol phospholipid hydrolysis, exhibited only partial desensitization to neurotensin. This result led to the observation that neurotensin was very rapidly degraded by intact N1E-115 cells. In experiments measuring the time course of [3H]neurotensin degradation, a minimum of six major tritiated products were found, with the breakdown peptides formed and the degree of proteolysis of [3H]neurotensin being dependent upon the length of incubation and the concentration of cells. Clone N1E-115 degraded [3H]neurotensin in an apparently sequential fashion; the primary initial cleavage of intact neurotensin was at the peptide bond between residues Arg8 and Arg9. Initial degradation peptides from the active carboxyl-terminal portion of neurotensin were more rapidly degraded, after formation, than were the peptides from the inactive amino-terminal half of neurotensin. The final two degradation products found were tyrosine, from the carboxyl-terminal portion of neurotensin, and an as yet unidentified peptide from the amino-terminal half of neurotensin. [3H]Neurotensin(8-13) was more rapidly hydrolyzed under identical conditions than was [3H]neurotensin itself. A combination of the protease inhibitors 1,10-phenanthroline and Z-Pro-Prolinal was able to inhibit almost completely the degradation of neurotensin by clone N1E-115.
小鼠神经母细胞瘤克隆N1E-115具有神经降压素受体，可介导细胞内环鸟苷酸的形成并刺激肌醇磷脂水解，该克隆对神经降压素仅表现出部分脱敏作用。这一结果导致观察到神经降压素在完整的N1E-115细胞中会非常迅速地降解。在测量[3H]神经降压素降解时间进程的实验中，发现至少有六种主要的氚化产物，[3H]神经降压素形成的降解肽段以及蛋白水解程度取决于孵育时间和细胞浓度。克隆N1E-115以一种明显有序的方式降解[3H]神经降压素；完整神经降压素的初始切割主要发生在残基Arg8和Arg9之间的肽键处。来自神经降压素活性羧基末端部分的初始降解肽段在形成后比来自神经降压素无活性氨基末端一半的肽段降解得更快。发现的最后两种降解产物是来自神经降压素羧基末端部分的酪氨酸和来自神经降压素氨基末端一半的一种尚未鉴定的肽段。在相同条件下，[3H]神经降压素(8-13)比[3H]神经降压素本身水解得更快。蛋白酶抑制剂1,10-菲咯啉和Z-脯氨酰-脯氨醛的组合能够几乎完全抑制克隆N1E-115对神经降压素的降解。
