The Yarrowia lipolytica Ribosomal Promoter pRPL25 as a Promising Object for Metabolic Engineering of Production Strains

Abstract

Promoters are crucial components of yeast production strains in metabolic engineering. Due to its high activity, the promoter of the RPL25 gene, which encodes the L25 protein of the large ribosomal subunit, is a promising object for investigation and optimization. Here, to investigate the Yarrowia lipolytica pRPL25 promoter structure, a randomized library of its sequences was generated in 100 nucleotide increments. The strength of the obtained promoters was evaluated using the green fluorescent protein as a reporter. The results showed that the minimum functional length of the studied pRPL25 derivatives was 199 bp, and the optimal functional length was 497 bp. When the sequence was shortened to 400 bp, the expression level of the reporter decreased significantly. Based on the above, we propose that the 400–500 bp region of pRPL25 comprises an upstream activating sequence (UAS). The pRPL25 promoter and its derivatives potentially can be used in bioengineering to create metabolic pathways with a precise level of gene expression. Strains constructed on their basis may be more efficient producers of target compounds compared to classical expression systems such as Saccharomyces cerevisiae.