Laboratoire de Biochimie des Protéines et Protéomique, UMR CNRS 7033, Université Paris 13, 74 rue Marcel Cachin, F-93017 Bobigny cedex, France.
Talanta. 2010 Feb 15;80(4):1576-85. doi: 10.1016/j.talanta.2009.07.016. Epub 2009 Jul 10.
The yeast Yarrowia lipolytica uses hydrophobic substrates, such as alkanes, fatty acids and oils, for its growth. It has developed a strategy for the use of such substrates, involving the production of hydrophobic binding structures called protrusions on the cell surface. These protrusions are resemble channels connecting the cell wall to the inside of the cell, and are probably involved in transport mechanisms that we do not yet fully understand. The complete genome of the haploid Y. lipolytica strain E150 (CLIB99) was sequenced in 2004 by the Génolevures Consortium. The availability of a complete genome sequence for this species has made it possible to carry out proteomic and other investigations, leading to the characterization of lipid bodies (LB) in terms of (i) their lipid composition, (ii) the major LB proteins, as identified by mass spectrometry, and (iii) differences in protein or lipid composition as a function of the carbon source used. Functional analyses would provide insight into the biological processes associated with these bodies and 2D BN/SDS-PAGE is a highly suitable method for the analysis of protein complexes. This report provides a first description of the analysis and identification of hydrophobic binding protein complexes in Y. lipolytica. For this purpose, we used 2D BN/SDS-PAGE for the separation of protein complexes and HPLC-chip-MS for protein identification. We separated and identified 40 protein complexes (11 heteromultimeric and 29 homomultimeric), providing insight into their function. This study represents a major step forward, as most previous studies identified proteins either on the basis of sequence similarity to proteins from other organisms (44% of the proteins identified in this study) or by prediction (50% of proteins identified in this study) alone.
酵母解脂耶氏酵母利用疏水性底物，如烷烃、脂肪酸和油脂，进行生长。它已经开发出一种利用这些底物的策略，涉及在细胞表面产生称为突起的疏水性结合结构。这些突起类似于连接细胞壁和细胞内部的通道，可能涉及我们尚未完全理解的运输机制。2004 年，由 Génolevures 联合体对单倍体 Y. lipolytica 菌株 E150 (CLIB99) 的全基因组进行了测序。该物种的完整基因组序列的可用性使得进行蛋白质组学和其他研究成为可能，从而对脂质体 (LB) 进行了描述，包括 (i) 其脂质组成、(ii) 通过质谱鉴定的主要 LB 蛋白，以及 (iii) 作为使用的碳源的函数的蛋白质或脂质组成的差异。功能分析将深入了解与这些体相关的生物学过程，而 2D BN/SDS-PAGE 是分析蛋白质复合物的高度合适的方法。本报告首次描述了 Y. lipolytica 中疏水性结合蛋白复合物的分析和鉴定。为此，我们使用 2D BN/SDS-PAGE 分离蛋白质复合物，使用 HPLC-chip-MS 鉴定蛋白质。我们分离和鉴定了 40 个蛋白质复合物（11 个异源多聚体和 29 个同源多聚体），深入了解了它们的功能。这项研究是向前迈出的重要一步，因为大多数以前的研究要么基于与其他生物体蛋白质的序列相似性来鉴定蛋白质（本研究中鉴定的蛋白质的 44%），要么仅基于预测（本研究中鉴定的蛋白质的 50%）。
