A simple generic method for optimizing membrane protein overexpression in Escherichia coli is still lacking. We have studied the physiological response of the widely used “Walker strains” C41(DE3) and C43(DE3), which are derived from BL21(DE3), to membrane protein overexpression. For unknown reasons, overexpression of many membrane proteins in these strains is hardly toxic, often resulting in high overexpression yields. By using a combination of physiological, proteomic, and genetic techniques we have shown that mutations in the lac UV5 promoter governing expression of T7 RNA polymerase are key to the improved membrane protein overexpression characteristics of the Walker strains. Based on this observation, we have engineered a derivative strain of E. coli BL21(DE3), termed Lemo21(DE3), in which the activity of the T7 RNA polymerase can be precisely controlled by its natural inhibitor T7 lysozyme (T7Lys). Lemo21(DE3) is tunable for membrane protein overexpression and conveniently allows optimizing overexpression of any given membrane protein by using only a single strain rather than a multitude of different strains. The generality and simplicity of our approach make it ideal for high-throughput applications.

We have investigated the conditions required for polar localization of the CheZ phosphatase by using a CheZ-green fluorescent protein fusion protein that, when expressed from a single gene in the chromosome, restored chemotaxis to a Δ cheZ strain. Localization was observed in wild-type, Δ cheZ , Δ cheYZ , and Δ cheRB cells but not in cells with cheA , cheW , or all chemoreceptor genes except aer deleted. Cells making only CheA-short (CheA _S ) or CheA lacking the P2 domain also retained normal localization, whereas cells producing only CheA-long or CheA missing the P1 and P2 domains did not. We conclude that CheZ localization requires the truncated C-terminal portion of the P1 domain present in CheA _S . Missense mutations targeting residues 83 through 120 of CheZ also abolished localization. Two of these mutations do not disrupt chemotaxis, indicating that they specifically prevent interaction with CheA _S while leaving other activities of CheZ intact.

Silver is well known for its antibacterial properties and low toxicity, and it is currently widely used both in the form of ions and of nanoparticles in many diverse products. One-dimensional silver nanowires (AgNWs) have the potential to further enhance the properties of nanosilver-containing products, since they appear to have higher antimicrobial efficacy and lower cytotoxicity. While they are widely used in optics and electronics, more studies are required in order to better understand their behavior in the biological environment and to be able to advance their application in uses such as wound healing, surface coating and drug delivery.