去泛素化酶USP14底物的鑒定及其相關(guān)功能的探究
發(fā)布時(shí)間：2018-12-07 07:30
【摘要】：去泛素化酶USP14是泛素特異性蛋白酶家族(ubiquitin-specific processing enzymes,UBPs/ubiquitin-specific proteases,USPs)中唯一一個(gè)與26S蛋白酶體相互作用的蛋白。其通過(guò)切除底物蛋白的泛素鏈來(lái)穩(wěn)定蛋白,從而影響信號(hào)通路、神經(jīng)系統(tǒng)發(fā)育和腫瘤發(fā)生發(fā)展等多種生理病理進(jìn)程。然而,USP14的底物以及其他生物功能一直未能得到很好的闡述,這嚴(yán)重阻礙了人們對(duì)于USP14在細(xì)胞生理進(jìn)程中作用機(jī)理的研究。我們通過(guò)定量蛋白質(zhì)組學(xué)手段,并輔以蛋白免疫沉淀和質(zhì)譜聯(lián)用(IP-MS)的分析方法,首次對(duì)USP14的調(diào)控網(wǎng)絡(luò)以及相互作用蛋白網(wǎng)絡(luò)進(jìn)行了研究。通過(guò)這些分析方法,我們得到了530個(gè)潛在相互作用蛋白,并從中選出8個(gè)USP14的潛在底物蛋白,FASN、CKB、ACADM、GNAS、IDH2、ABLIM1、MYO1E和CTSB,這些蛋白在能量代謝以及細(xì)胞命運(yùn)中均起著重要作用。此外,我們通過(guò)泛素化肽段的富集鑒定到了12178個(gè)賴氨酸泛素化位點(diǎn),其中CKB、FASN、UBB和MYO1E的部分泛素化位點(diǎn)均有一定上調(diào),進(jìn)一步暗示這些蛋白是USP14的底物蛋白。同時(shí),我們結(jié)合蛋白相互作用譜和泛素化修飾譜數(shù)據(jù)篩選了一些其他潛在蛋白底物。這些結(jié)果對(duì)于進(jìn)一步了解USP14的作用機(jī)理以及生物功能均有重要意義。
[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.
【學(xué)位授予單位】：中國(guó)科學(xué)院上海藥物研究所
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2016
【分類(lèi)號(hào)】：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.
【學(xué)位授予單位】：中國(guó)科學(xué)院上海藥物研究所
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2016
【分類(lèi)號(hào)】：R927
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