Laboratory of Computational Systems Biotechnology, EPFL, CH-1015 Lausanne, Switzerland.
Biotechnol Bioeng. 2012 Aug;109(8):2127-33. doi: 10.1002/bit.24463. Epub 2012 Feb 24.
Despite the establishment of design principles to optimize codon choice for heterologous expression vector design, the relationship between codon sequence and final protein yield remains poorly understood. In this work, we present a computational framework for the identification of a set of mutant codon sequences for optimized heterologous protein production, which uses a codon-sequence mechanistic model of protein synthesis. Through a sensitivity analysis on the optimal steady state configuration of protein synthesis we are able to identify the set of codons, that are the most rate limiting with respect to steady state protein synthesis rate, and we replace them with synonymous codons recognized by charged tRNAs more efficient for translation, so that the resulting codon-elongation rate is higher. Repeating this procedure, we iteratively optimize the codon sequence for higher protein synthesis rate taking into account multiple constraints of various types. We determine a small set of optimized synonymous codon sequences that are very close to each other in sequence space, but they have an impact on properties such as ribosomal utilization or secondary structure. This limited number of sequences can then be offered for further experimental study. Overall, the proposed method is very valuable in understanding the effects of the different properties of mRNA sequences on the final protein yield in heterologous protein production and it can find applications in synthetic biology and biotechnology.
尽管已经建立了设计原则来优化用于异源表达载体设计的密码子选择，但密码子序列与最终蛋白质产量之间的关系仍未得到很好的理解。在这项工作中，我们提出了一种用于识别一组用于优化异源蛋白质生产的突变密码子序列的计算框架，该框架使用蛋白质合成的密码子序列机制模型。通过对蛋白质合成的最优稳态配置进行敏感性分析，我们能够确定一组密码子，这些密码子对于稳态蛋白质合成速率的限制最大，然后用翻译效率更高的负载 tRNA 识别的同义密码子替换它们，从而使生成的密码子延伸速率更高。通过重复此过程，我们考虑到各种类型的多个约束，迭代优化密码子序列以提高蛋白质合成速率。我们确定了一小组优化的同义密码子序列，它们在序列空间中非常接近，但它们对核糖体利用或二级结构等性质有影响。然后，可以提供这些有限数量的序列供进一步的实验研究。总的来说，该方法对于理解 mRNA 序列的不同性质对异源蛋白质生产中最终蛋白质产量的影响非常有价值，并且可以在合成生物学和生物技术中找到应用。
