去泛素化酶USP14底物的鉴定及其相关功能的探究
发布时间：2018-12-07 07:30
【摘要】：去泛素化酶USP14是泛素特异性蛋白酶家族(ubiquitin-specific processing enzymes,UBPs/ubiquitin-specific proteases,USPs)中唯一一个与26S蛋白酶体相互作用的蛋白。其通过切除底物蛋白的泛素链来稳定蛋白,从而影响信号通路、神经系统发育和肿瘤发生发展等多种生理病理进程。然而,USP14的底物以及其他生物功能一直未能得到很好的阐述,这严重阻碍了人们对于USP14在细胞生理进程中作用机理的研究。我们通过定量蛋白质组学手段,并辅以蛋白免疫沉淀和质谱联用(IP-MS)的分析方法,首次对USP14的调控网络以及相互作用蛋白网络进行了研究。通过这些分析方法,我们得到了530个潜在相互作用蛋白,并从中选出8个USP14的潜在底物蛋白,FASN、CKB、ACADM、GNAS、IDH2、ABLIM1、MYO1E和CTSB,这些蛋白在能量代谢以及细胞命运中均起着重要作用。此外,我们通过泛素化肽段的富集鉴定到了12178个赖氨酸泛素化位点,其中CKB、FASN、UBB和MYO1E的部分泛素化位点均有一定上调,进一步暗示这些蛋白是USP14的底物蛋白。同时,我们结合蛋白相互作用谱和泛素化修饰谱数据筛选了一些其他潜在蛋白底物。这些结果对于进一步了解USP14的作用机理以及生物功能均有重要意义。
[Abstract]:USP14 is the only protein in the ubiquitin specific protease family (ubiquitin-specific processing enzymes,UBPs/ubiquitin-specific proteases,USPs) that interacts with 26s proteasome. It can stabilize the protein by removing the ubiquitin chain of substrate protein, thus affecting many physiological and pathological processes, such as signal pathway, nervous system development and tumorigenesis and development. However, the substrate and other biological functions of USP14 have not been well elucidated, which seriously hinders the study of the mechanism of USP14 in cellular physiological process. The regulatory and interacting protein networks of USP14 were studied for the first time by quantitative proteomics and protein immunoprecipitation and mass spectrometry (IP-MS). Through these analysis methods, we obtained 530 potential interaction proteins, from which 8 USP14 potential substrate proteins, FASN,CKB,ACADM,GNAS,IDH2,ABLIM1,MYO1E and CTSB, were selected. These proteins play an important role in energy metabolism and cell fate. In addition, 12178 Lysine ubiquitin sites were identified by enrichment of Ubiquitin peptides, in which some Ubiquitin sites of CKB,FASN,UBB and MYO1E were up-regulated, which further suggested that these proteins were substrates of USP14. At the same time, we selected some other potential protein substrates by binding protein interaction spectra and ubiquitin modification spectra data. These results are important for further understanding the mechanism and biological function of USP14.
【学位授予单位】：中国科学院上海药物研究所
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R927
[Abstract]:USP14 is the only protein in the ubiquitin specific protease family (ubiquitin-specific processing enzymes,UBPs/ubiquitin-specific proteases,USPs) that interacts with 26s proteasome. It can stabilize the protein by removing the ubiquitin chain of substrate protein, thus affecting many physiological and pathological processes, such as signal pathway, nervous system development and tumorigenesis and development. However, the substrate and other biological functions of USP14 have not been well elucidated, which seriously hinders the study of the mechanism of USP14 in cellular physiological process. The regulatory and interacting protein networks of USP14 were studied for the first time by quantitative proteomics and protein immunoprecipitation and mass spectrometry (IP-MS). Through these analysis methods, we obtained 530 potential interaction proteins, from which 8 USP14 potential substrate proteins, FASN,CKB,ACADM,GNAS,IDH2,ABLIM1,MYO1E and CTSB, were selected. These proteins play an important role in energy metabolism and cell fate. In addition, 12178 Lysine ubiquitin sites were identified by enrichment of Ubiquitin peptides, in which some Ubiquitin sites of CKB,FASN,UBB and MYO1E were up-regulated, which further suggested that these proteins were substrates of USP14. At the same time, we selected some other potential protein substrates by binding protein interaction spectra and ubiquitin modification spectra data. These results are important for further understanding the mechanism and biological function of USP14.
【学位授予单位】：中国科学院上海药物研究所
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R927
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