由于细胞体内核糖体蛋白的多样性及核糖体结构的复杂性，人们较少在核糖体蛋白及核糖体翻译水平上研究细胞外源蛋白合成调控机制。毕赤酵母为重要外源蛋白表达工业菌株，我们关于毕赤酵母外源蛋白表达及蛋白质组分析中发现：重组毕赤酵母核糖体蛋白下调与细胞生长及外源蛋白过量表达密切相关。本课题拟采用iTRAQ LC-MS/MS分析重组毕赤酵母核糖体蛋白及能量代谢途径的差异表达，结合基于Ribo-seq定量分析核糖体蛋白差异表达对核糖体翻译效率的影响，尤其是对外源蛋白mRNA翻译的影响，分析多核糖体构成与分布变化，研究核糖体蛋白对毕赤酵母细胞生长及外源蛋白合成的调控规律。并通过对表达下调的核糖体蛋白基因缺失构建毕赤酵母突变菌株，分析核糖体蛋白对细胞核糖体翻译、内质网胁迫耐受及蛋白合成分泌等功能的影响，深入探讨核糖体蛋白对毕赤酵母细胞生长及外源蛋白合成的分子调控机制，为毕赤酵母核糖体优化与定制提供基础。

Work published in the past year has shown that changes to the ribosome core can affect the mechanism of translation, which potentially leads to specific changes in the relative efficiencies with which different proteins are made. However, few of the translational regulation of heterologous protein synthesis were reported because of the diversity of ribosomal proteins (RPs) and complexity of ribosome structure. The methylotrophic yeast Pichia pastoris is a powerful eukaryotic platform for the production of heterologous protein. Recent publication of the Pichia pastoris genome has facilitated strain development toward biopharmaceutical and environmental science applications, developed the study on transcriptomics and proteomics. On quantitative iTRAQ LC-MS/MS proteomics of the cellular response to heterologous protein overexpression and the regulation of HAC1 in Pichia pastoris, we observed that over 30 of RPs exited the different expression and accompanied with the slower of cells growth. This result is hypothesized that the different expression of RPs would impact the ribosome structure and translational efficiency, especial translation of heterologous protein. To investigate functional characterization of RPs from Pichia pastoris and regulation on heterologous protein synthesis, the ribosome structure with polyribosome analysis and translational efficiency using ribosome-sequence will be carried on. The further research will be developed to RPs gene deletion strains to validate its ribosome translational function and non-ribosome function,containing the protection of ER stress,enhancing scretion of heterologous protein, and so on. It will expand the database of RPs genes and the investigation of ribosome engineering.