Overexpression of Mouse Estrogen Receptor-β Decreases but Its Transactivation and Ligand Binding Domains Increase the Growth Characteristics of E. coli

Abstract

Escherichia coli is one of the most common and widely used prokaryotic hosts for the expression of recombinant proteins. The overexpression of recombinant proteins occasionally increases bacterial growth but sometimes reduces it and becomes lethal to the host cells. Here, we report the overexpression of mouse ER-β and its domains in the prokaryotic expression system and its opposite effect on the growth characteristics of E. coli. ER-β protein was immunologically detected as a 53 kDa his-tag protein in the pellet of the bacterial lysate. Its overexpression, as reflected by the total protein content and expression pattern, resulted in the decrease of bacterial growth. However, the overexpression of ER-β transactivation domain (TAD) using pIVEX and ligand binding domain (LBD) using pRSETA in E. coli BL21 (DE3) show opposite pattern. TAD was immunologically detected as 20 kDa and LBD as 22 kDa protein in the supernatant of the bacterial lysate and their overexpression increased the bacterial growth.

ER<i>β</i> interacting protein ERAP 140 shows age dependent alteration in its interaction and expression in male mouse brain

“…In continuation, we reported that mouse ERLBD interacts with PELP1, RIP140, PGC1α, BAF60 and ADA3 [33,34], and ERTAD with metastasis associated protein (MTA) 1 and p68 RNA helicase [35], and these coregulators vary in their levels of interaction and expression during aging of mouse brain. For the identification ERβ interacting proteins in mouse brain, purified [36] and functionally active [37] ERβ was used. Recently, we reported age and sex dependent interaction of AIB1 with ERβTAD and its expression in mouse brain [14].…”

Cloning, improved expression and purification of invasion plasmid antigen D (IpaD): an effector protein of enteroinvasive Escherichia coli (EIEC)

“…2E). Additionally, higher IPTG concentration had undesirable effects on cell growth and soluble protein yield which may be attributed to the increase of metabolic load because target protein overexpression by bacterial cells and results in interrupted metabolic activity and overall decrease of protein yield (Paramanik & Thakur 2011). This observation is supported by previous reports recommending the use of low IPTG concentration for induction owing to the following reasons i) higher concentrations are toxic to cell, ii) expensive, iii) potential problem of product insolubility, iv) It induces growth inhibition and cell lysis (Donovan et al 1996;Volontè et al 2008).…”

AIB1 shows variation in interaction with ERβTAD and expression as a function of age in mouse brain

“…Subsequently, the bacterial cells were collected after centrifugation at 3,0009g for 10 min at 4°C followed by incubation in lysis buffer (20 mM Tris HCl pH 8.0, 10 mM imidazole, 150 mM NaCl, 0.2% Triton X-100 and 2 mM b-mercaptoethanol). Then, the cell lysate was processed for the purification of recombinant proteins (Paramanik and Thakur 2010a).…”