Castro Leandro M, Cavalcanti Diogo M L P, Araujo Christiane B, Rioli Vanessa, Icimoto Marcelo Y, Gozzo Fábio C, Juliano Maria, Juliano Luiz, Oliveira Vitor, Ferro Emer S
Department of Pharmacology, Support Center for Research in Proteolysis and Cell Signaling (NAPPS), University of São Paulo, 05508-000 SP, Brazil; Department of Biophysics, Federal University of São Paulo, 04039-032 SP, Brazil.
Department of Pharmacology, Support Center for Research in Proteolysis and Cell Signaling (NAPPS), University of São Paulo, 05508-000 SP, Brazil.
J Proteomics. 2014 Dec 5;111:238-48. doi: 10.1016/j.jprot.2014.03.043. Epub 2014 Apr 13.
A large number of intracellular peptides are constantly produced following protein degradation by the proteasome. A few of these peptides function in cell signaling and regulate protein-protein interactions. Neurolysin (Nln) is a structurally defined and biochemically well-characterized endooligopeptidase, and its subcellular distribution and biological activity in the vertebrate brain have been previously investigated. However, the contribution of Nln to peptide metabolism in vivo is poorly understood. In this study, we used quantitative mass spectrometry to investigate the brain peptidome of Nln-knockout mice. An additional in vitro digestion assay with recombinant Nln was also performed to confirm the identification of the substrates and/or products of Nln. Altogether, the data presented suggest that Nln is a key enzyme in the in vivo degradation of only a few peptides derived from proenkephalin, such as Met-enkephalin and octapeptide. Nln was found to have only a minor contribution to the intracellular peptide metabolism in the entire mouse brain. However, further studies appear necessary to investigate the contribution of Nln to the peptide metabolism in specific areas of the murine brain.
Neurolysin was first identified in the synaptic membranes of the rat brain in the middle 80's by Frederic Checler and colleagues. Neurolysin was well characterized biochemically, and its brain distribution has been confirmed by immunohistochemical methods. The neurolysin contribution to the central and peripheral neurotensin-mediated functions in vivo has been delineated through inhibitor-based pharmacological approaches, but its genuine contribution to the physiological inactivation of neuropeptides remains to be firmly established. As a result, the main significance of this work is the first characterization of the brain peptidome of the neurolysin-knockout mouse. This article is part of a Special Issue entitled: Proteomics, mass spectrometry and peptidomics, Cancun 2013. Guest Editors: César López-Camarillo, Victoria Pando-Robles and Bronwyn Jane Barkla.
蛋白酶体降解蛋白质后会持续产生大量细胞内肽段。其中少数肽段参与细胞信号传导并调节蛋白质 - 蛋白质相互作用。神经溶素（Nln）是一种结构明确且生化特性良好的内肽酶，此前已对其在脊椎动物大脑中的亚细胞分布和生物学活性进行过研究。然而，Nln在体内对肽代谢的贡献仍知之甚少。在本研究中，我们使用定量质谱法研究Nln基因敲除小鼠的脑肽组。还进行了一项额外的重组Nln体外消化试验，以确认Nln底物和/或产物的鉴定。总体而言，所呈现的数据表明，Nln是体内仅少数源自脑啡肽原的肽段（如甲硫氨酸脑啡肽和八肽）降解的关键酶。发现Nln对整个小鼠脑内的细胞内肽代谢贡献较小。然而，似乎有必要进一步研究Nln对小鼠脑特定区域肽代谢的贡献。
神经溶素最早于80年代中期由弗雷德里克·谢克莱尔及其同事在大鼠脑的突触膜中发现。神经溶素在生化方面已得到充分表征，其在脑中的分布已通过免疫组织化学方法得到证实。通过基于抑制剂的药理学方法已经阐明了神经溶素对体内中枢和外周神经降压素介导功能的贡献，但其对神经肽生理失活的真正贡献仍有待确定。因此，这项工作的主要意义在于首次对神经溶素基因敲除小鼠的脑肽组进行表征。本文是题为：蛋白质组学、质谱和肽组学，2013年坎昆的特刊的一部分。客座编辑：塞萨尔·洛佩斯 - 卡马里略、维多利亚·潘多 - 罗夫莱斯和布朗温·简·巴克勒。
