Abstract
The Gram-negative bacterium E. coli is the host of choice for a multitude of used recombinant proteins. Generally, cultivation is easy, media are cheap, and a high product titer can be obtained. However, harsh induction procedures using isopropyl β-d-1 thiogalactopyranoside as inducer are often referred to cause stress reactions, leading to a phenomenon known as “metabolic” or “product burden”. These high expressions of recombinant proteins mainly result in decreased growth rates and cell lysis at elevated induction times. Therefore, approaches tend to use “soft” or “tunable” induction with lactose and reduce the stress level of the production host. The usage of glucose as energy source in combination with lactose as induction reagent causes catabolite repression effects on lactose uptake kinetics and as a consequence reduced product titer. Glycerol—as an alternative carbon source—is already known to have positive impact on product formation when coupled with glucose and lactose in auto-induction systems, and has been referred to show no signs of repression when cultivated with lactose concomitantly. In recent research activities, the impact of different products on the lactose uptake using glucose as carbon source was highlighted, and a mechanistic model for glucose-lactose induction systems showed correlations between specific substrate uptake rate for glucose or glycerol (qs,C) and the maximum specific lactose uptake rate (qs,lac,max). In this study, we investigated the mechanistic of glycerol uptake when using the inducer lactose. We were able to show that a product-producing strain has significantly higher inducer uptake rates when being compared to a non-producer strain. Additionally, it was shown that glycerol has beneficial effects on viability of cells and on productivity of the recombinant protein compared to glucose.
Highlights
The Gram-negative bacterium E. coli is the expression host of choice for the production of 30%to 40% of recombinant drugs in industry [1,2]
As tunable protein production is commonly applied in industry nowadays, mixed-feed systems using either isopropyl β-D-1 thiogalactopyranoside (IPTG) [21] or lactose [13,22,23] as inducer did result in higher product yields when compared to other inducer supplies [24]
As lactose can be metabolized in E. coli, it does not stress the cells as much as IPTG [26]
Summary
The Gram-negative bacterium E. coli is the expression host of choice for the production of 30%to 40% of recombinant drugs in industry [1,2]. As tunable protein production is commonly applied in industry nowadays, mixed-feed systems using either IPTG [21] or lactose [13,22,23] as inducer did result in higher product yields when compared to other inducer supplies [24]. Soft induction performed with lactose shows promising results [13,23,25]. As lactose can be metabolized in E. coli, it does not stress the cells as much as IPTG [26]. For the production of soluble proteins, induction with lactose usually is preferred [3], but it has been shown that lactose shows promising results for Inclusion Bodies (IBs) and products located in the periplasm [3,27]
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