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首页> 外文期刊>Virology Journal >Proteome analysis of vaccinia virus IHD-W-infected HEK 293 cells with 2-dimensional gel electrophoresis and MALDI-PSD-TOF MS of on solid phase support N-terminally sulfonated peptides
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Proteome analysis of vaccinia virus IHD-W-infected HEK 293 cells with 2-dimensional gel electrophoresis and MALDI-PSD-TOF MS of on solid phase support N-terminally sulfonated peptides

机译：固相支持N末端磺化肽的二维凝胶电泳和MALDI-PSD-TOF MS对牛痘病毒IHD-W感染的HEK 293细胞进行蛋白质组分析

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Background Despite the successful eradication of smallpox by the WHO-led vaccination programme, pox virus infections remain a considerable health threat. The possible use of smallpox as a bioterrorism agent as well as the continuous occurrence of zoonotic pox virus infections document the relevance to deepen the understanding for virus host interactions. Since the permissiveness of pox infections is independent of hosts surface receptors, but correlates with the ability of the virus to infiltrate the antiviral host response, it directly depends on the hosts proteome set. In this report the proteome of HEK293 cells infected with Vaccinia Virus strain IHD-W was analyzed by 2-dimensional gel electrophoresis and MALDI-PSD-TOF MS in a bottom-up approach. Results The cellular and viral proteomes of VACV IHD-W infected HEK293 cells, UV-inactivated VACV IHD-W-treated as well as non-infected cells were compared. Derivatization of peptides with 4-sulfophenyl isothiocyanate (SPITC) carried out on ZipTipμ-C18 columns enabled protein identification via the peptides' primary sequence, providing improved s ratios as well as signal intensities of the PSD spectra. The expression of more than 24 human proteins was modulated by the viral infection. Effects of UV-inactivated and infectious viruses on the hosts' proteome concerning energy metabolism and proteins associated with gene expression and protein-biosynthesis were quite similar. These effects might therefore be attributed to virus entry and virion proteins. However, the modulation of proteins involved in apoptosis was clearly correlated to infectious viruses. Conclusions The proteome analysis of infected cells provides insight into apoptosis modulation, regulation of cellular gene expression and the regulation of energy metabolism. The confidence of protein identifications was clearly improved by the peptides' derivatization with SPITC on a solid phase support. Some of the identified proteins have not been described in the context of poxvirus infections before and need to be further characterised to identify their meaning for apoptosis modulation and pathogenesis.
机译：背景技术尽管世卫组织领导的疫苗接种计划成功根除了天花，但痘病毒感染仍然是相当大的健康威胁。天花可能被用作生物恐怖剂以及人畜共患痘病毒感染的持续发生证明了加深对病毒宿主相互作用的认识。由于痘病毒的容许性与宿主表面受体无关，但与病毒渗透抗病毒宿主反应的能力相关，因此它直接取决于宿主蛋白质组。在该报告中，通过二维凝胶电泳和MALDI-PSD-TOF MS，以自下而上的方法分析了感染了痘苗病毒IHD-W的HEK293细胞的蛋白质组。结果比较了VACV IHD-W感染的HEK293细胞，紫外线灭活的VACV IHD-W处理以及未感染的细胞的细胞和病毒蛋白质组。在ZipTipμ-C18色谱柱上进行的4-硫代苯基异硫氰酸酯（SPITC）衍生化肽能够通过肽的一级序列进行蛋白质鉴定，从而提高了s / n比以及PSD光谱的信号强度。病毒感染调节了24种以上人类蛋白质的表达。紫外线灭活和感染性病毒对宿主蛋白质组的能量代谢以及与基因表达和蛋白质生物合成相关的蛋白质的影响非常相似。因此，这些影响可能归因于病毒进入和病毒粒子蛋白。然而，参与凋亡的蛋白质的调节显然与传染性病毒有关。结论感染细胞的蛋白质组分析可深入了解细胞凋亡的调控，细胞基因表达的调控和能量代谢的调控。通过肽在固相支持物上的SPITC衍生化，明显提高了蛋白质鉴定的可信度。以前在痘病毒感染的背景下尚未描述某些鉴定出的蛋白质，需要进一步表征以鉴定其对细胞凋亡调节和发病机理的意义。

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