Surface Display of Bacterial Metallothioneins and a Chitin Binding Domain on Escherichia coli Increase Cadmium Adsorption and Cell Immobilization

Abstract

To increase the level of adsorption of cadmium ions to the surface of Escherichia coli, we fused cyanobacterial metallothioneins, SmtA (from Synechococcus elongatus PCC 3601) and MtnA (from Synechococcus vulcanus) to the E. coli cell surface using a Lpp'-OmpA-based display system. E. coli strains expressing Lpp'-OmpA-SmtA-linker-ChBD (chitin-binding domain from Bacillus pumillus SG2 chitinase S; chiS) and Lpp'-OmpA-MtnA-linker-ChBD on their surface adsorbed more cadmium compared to the E. coli cells expressing only the Lpp'-OmpA-linker-ChBD hybrid. These constructs also were bound to chitin through their chitin-binding domain, allowing them to be immobilized on a chitin matrix. We assessed surface presentation of Lpp'-OmpA-SmtA-linker-ChBD, Lpp'-OmpA-MtnA-linker-ChBD, and Lpp'-OmpA-linker-ChBD using immunostaining. The Lpp'-OmpA-SmtA-linker-ChBD chimera adsorbed metal and was bound to chitin with the highest efficiency compared to the other chimeras, suggesting that it is an effective bioadsorbent. This is the first example of coupling metal adsorption with cell immobilization using a whole-cell bioadsorbent.

Molecular dynamics simulation and experimental study of the surface-display of SPA protein via Lpp-OmpA system for screening of IgG

“…The e ciency of surface display systems and the correct and e cient protein folding and its stability is highly related to the speci cations of the carrier protein, passenger protein, and fusion method Barrett ., et al 2019). LPP-ompA is an e cient surface display system that has been developed and applied for various applications (Fasehee ., et al 2018;Rigi ., et al 2014 ;Tafakori., et al 2012). In this research project using Lpp'-OmpA as an SPA anchor , in the initial design we performed physico-chemical and structural studies on chimeric protein using molecular dynamics tools to ensure the strength and stability of this new structure on the cell surface.…”

“…Display of heterologous proteins on the bacterial surface has been demonstrated as a multi-strategy approach to develop an e cient vaccine for S. aureus development (Kalyanasundram et al 2015), screening of antibody libraries (Cavallari 2017), development of whole-cell bioadsorbents (Tafakori et al 2012), and biosensors (Furst et al 2017). Chimeric protein system of the Lpp′-OmpA is used as an anchor and loads heterologous proteins onto the Gram-negative bacterial surface Georgiou et al 1996).…”

Molecular dynamics simulation and experimental study of the surface-display of SPA protein via Lpp-OmpA system for screening of IgG

“…Display of heterologous proteins on the bacterial surface has been demonstrated as a multi-strategy approach to develop an e cient vaccine for S. aureus development (Kalyanasundram et al, 2015), screening of antibody libraries (Cavallari, 2017), development of whole-cell bioadsorbents (Tafakori et al, 2012), and biosensors (Furst et al, 2017). Chimeric protein system of the Lpp′-OmpA is used as an anchor and loads heterologous proteins onto the Gram-negative bacterial surface (Yang et al, Georgiou et al, 1996).…”

Engineering E. coli cell surface in order to develop a one-step purification method for recombinant proteins

“…Bacterial surface display of heterologous proteins has become an attractive strategy for a wide variety of applications like screening of antibody libraries (Cavallari 2017 ), as a vaccine delivery vehicle (Kalyanasundram et al 2015 ), production of cellular adsorbents (Tafakori et al 2012 ), and development of cellular biosensors (Liang et al 2015 ). In this process, a coding sequence for the passenger protein is fused to the gene encoding a bacterial surface protein.…”