Team:PennState/MedalChecklist

Diauxie Elimination

Introduction

The System

Strategies

Progress

Conclusions

Parts

References

NHR Biosensors

NHR Introduction

Phthalate Biosensor

BPA Biosensor

A common framework for gene expression at bacterial catabolic operons involves dual regulation by a global regulatory protein and a catabolite-specific regulator (e.g., AraC in the case of expression from promoter PBAD). In E. coli, the cAMP-receptor protein (CRP) acts as a global regulator in which the cAMP-CRP complex typically increases transcription at catabolic promoters in the absence of the “preferred” catabolite glucose. The result is a phenomenon known as diauxie, in which glucose is preferentially utilized in the presence of other sugars, since expression of catabolic pathways for the other sugars is not fully induced. A consequence of this dual control mechanism is that many bacterial promoters commonly used in biotechnology require the absence of glucose for full transcription activation (e.g., the lac and araBAD promoters).

In wild-type E. coli strains, the promoters controlling expression of genes responsible for xylose transport and metabolism are regulated by CRP and the xylose-inducible protein XylR. Our goal in this project is to create and characterize a xylose-inducible but glucose-insensitive gene expression system. This would functionally eliminate a diauxie-type phenotype relating to induction of gene expression from this promoter. In addition to creating a valuable new tool for the part registry, this project has useful applications for biochemical and bioenergy production. Cellulosic biomass feedstocks targeted for biofuel production or other value-added products contain large percentages of glucose and xylose. In industrial fermentations, cells grown on sugars from cellulosic biomass normally consume glucose as their first carbon source. Then, depending on the process, cells either change gene expression to utilize xylose, or the cells and leftover sugars are removed as waste. Both situations lead to inefficiency in production, especially if a continuous growth process is desired. Growing cells on multiple sugars results in a lag time as the cells switch from glucose to xylose metabolism, which complicates and delays the overall process. The gene expression system we are creating could aid in the simultaneous fermentation of mixed sugars, and would have practical applications during the conversion of biomass to ethanol, and for other processes using bacteria for fermentation of low-cost sugar mixtures.