Genetic Engineering of Caulobacter crescentus for Removal of Cadmium from Water

Abstract

Hexa-histidine (6His) peptide was inserted to a permissive site of the surface layer (S-layer) protein RsaA of Caulobacter crescentus. The recombinant strain JS4022/p723-6H, expressing RsaA-6His fusion protein was examined for its ability to sequester Cd(II) from the bacterial growth medium. When mixed with 1 ppm CdCl(2), JS4022/p723-6H removed 94.3 approximately 99.9% of the Cd(II), whereas the control strain removed only 11.4 approximately 37.0%, depending on experimental conditions. The effective contact time of the cells and Cd(II) was as short as 15 min. When higher concentrations of CdCl(2) were tested, JS4022/p723-6H consistently demonstrated enhanced binding capacity over the control strain. At 15 ppm of Cd(II), each gram of JS4022/p723-6H dry cells retrieved 16.0 mg of Cd(II), comparing to 11.6 mg g(-1) achieved by the control strain. This work provides a potential cost-effective solution toward bioremediation of heavy metals from aqueous systems.

Self-immobilization of Recombinant Caulobacter crescentus and Its Application in Removal of Cadmium from Water

“…Recombinant C. crescentus strain JS4022/p723-6H expressing RsaA-6His fusion proteins was constructed as described previously [10]. Strain JS4022/p4ArsaA(723Δ)GSCCΔ expressing RsaA with a cloning linker at its 723-amino-acid residual position [27] was used as a control for cadmium binding assays.…”

“…The efficacy of using biomass from plants [3,4], animals [5], and microbes [6][7][8][9] to sequester heavy metals is well documented. One popular approach is to incorporate heavy-metal-binding peptides such as polyhistidines [10,11], metallothionein [12,13], and synthetic phytochelatins [14] to the cell surface of microbes through genetic manipulations and to use the constructed strain as whole-cell adsorbents for heavy metals. Display of peptides that have high affinity to heavy metals at the bacterial cell surface minimizes uptake of metals across cell membranes, thereby alleviating toxicity to host cells.…”

“…Due to its high copy number, outermost location, and readiness for genetic manipulation, RsaA has become a popular carrier for display of foreign peptides on the surface of Caulobacter cells. In a previous study, we inserted hexahistidine (6His) polypeptide to a permissive site of the surface layer (S-layer) protein RsaA of Caulobacter [10]. The RsaA-6His fusion protein was successfully expressed to the outer surface of the recombinant strain JS4022/p723-6H and was able to specifically retrieve Cd(II) from solutions in the presence of an excess of other divalent ions, such as calcium and magnesium.…”

Bioremediation of soluble heavy metals with recombinantCaulobacter crescentus

“…We have reported the construction of a recombinant C. crescentus strain JS4022/ p723-6H with hexahistidine peptides (6His) inserted at a permissive site of the S-layer protein RsaA. 9 Because the engineered S-layer fusion protein RsaA-6His is displayed at the cell surface, strain JS4022/p723-6H can be directly used as a whole cell adsorbent for sequestration of dissolved heavy metals from water. When an appropriate amount of bacterial biomass is used, JS4022/p723-6H can remove 97∼99.9% of the total dissolved cadmium, as opposed to less than 20% achieved by the control strain JS4022/ p4ArsaA(723∆)GSCC∆, which expresses RsaA with a cloning linker inserted at the same permissive site.…”

“…Dr. John Smit's group at the University of British Columbia has identified materials that could hold up to nine times as many Caulobacter cells as glass surfaces within an hour (personal communication). When necessary, a biosupporting unit may be biofilms loses its stability in high concentrations of detergent, such as 0.1% (w/v) sodium dodecyl sulfate, 9 which poses a restriction on the types of waste water that can be treated with such systems. On the other hand, detergent wash can be used as a convenient way to eradicate the biofilms when needed.…”