Focused proteomic analysis of functional network for peptidyl prolyl cis/trans isomerases

肽基脯氨酰顺式/反式异构酶（PPlases）催化脯氨酸之前的肽键旋转，这是一个限制新合成蛋白质折叠速率的步骤。PPase还具有结合许多蛋白质并充当伴侣的能力;因此，它们被认为调节细胞中蛋白质的折叠、组装和运输，并控制蛋白质的活性。PPase是免疫抑制药物环孢菌素A（CsA）和FK 506最常见的靶点，它们分别与亲环蛋白（CyP）和FK 506结合蛋白（FKBP）结合并抑制其同源PPase活性。CyP和FKBP都是普遍存在的，从细菌到人都高度表达和保守。PPlases的第三个家族，即小蛋白，既不是CsA也不是FK 506的靶点，但也是从细菌到人类的保守蛋白。虽然CyP、FKBP和parvulin的同源物在几乎所有的生物中都已被鉴定，但这些同源物的细胞功能仍有待进一步研究。在这项研究中，我们系统地分析了23种不同的人类PPlase，包括9 CyP，11 FKBP，和3 parvulin同系物的潜在底物，通过使用蛋白质组学方法，包括表位标记的PPlase的表达，在存在或不存在的PPlase抑制剂的亲和纯化，基于质谱的蛋白质鉴定，和数据库搜索。我们鉴定了三种PPlases的约200种潜在底物，并构建了PPlases的功能蛋白网络。该网络包括在翻译、细胞周期、增殖、转录、应激反应、DNA/蛋白质代谢、RNA加工、蛋白质运输和核糖体生物合成中起作用的蛋白质。我们进一步研究了PPlases在核糖体生物合成中的作用，发现parvulin 14、FKBP 25和CyPB可能在人类核糖体生物合成的不同阶段发挥作用。

Peptidyl prolyl cis/trans isomerases (PPlases) catalyze the rotation about the peptide bond preceding proline, a step that can be rate-limiting for the folding of newly synthesized proteins. PPlases also have the ability to bind many proteins and to act as chaperones; thus they are believed to regulate folding, assembly and trafficking, and controlling activity of proteins in the cell. PPlases are most familiar as the targets of the immunosuppressive drugs, cyclosporin A (CsA) and FK506, which bind, respectively, to cyclophilin (CyP) and FK506-binding protein (FKBP) and inhibit their cognate PPlaseactivities. Both CyP and FKBP are ubiquitous, highly expressed and conserved from bacteria to human. The third family of PPlases, parvulins, is the target of neither CsA nor FK506, but is also conserved from bacteria to human. Although a number of CyP, FKBP and parvulin homologs have been identified in almost all organisms, the cellular functions of most of those homologs remain to be exploded. In this study, we analyzed systematically potential substrate for 23 different human PPlases including 9 CyP, 11 FKBP, and 3 parvulin homologs by using proteomic approaches that include expression of epitope-tagged PPlase, affinity purification in the presence or absence of PPlase inhibitor, mass-spectrometry based protein identification, and database searching. We identified about 200 potential substrates for the three types of PPlases and constructed a functional protein network for the PPlases. The network included proteins that have roles in translation, cell cycle, proliferation, transcription, stress-responses, DNA/protein metabolism, RNA processing, protein trafficking, and ribosome biogenesis. We investigated further the involvement of PPlases in ribosome biogenesis, and found that parvulin 14, FKBP25, and CyPB have probably roles in different stages of human ribosome biogenesis.