Efficient purification protocol for bioengineering allophycocyanin trimer with N-terminus Histag

Abstract EN

Allophycocyanin plays a key role for the photon energy transfer from the phycobilisome to reaction center chlorophylls with high efficiency in cyanobacteria.

Previously, the high soluble self-assembled bioengineering allophycocyanin trimer with N-terminus polyhistidine from Synechocystis sp.

PCC 6803 had been successfully recombined and expressed in Escherichia coli strain.

The standard protocol with immobilized metal-ion affinity chromatography with chelating transition metal ion (Ni2+) was used to purify the recombinant protein.

Extensive optimization works were performed to obtain the desired protocol for high efficiency, low disassociation, simplicity and fitting for large-scale purification.

In this study, a 33 full factorial response surface methodology was employed to optimize the varied factors such as pH of potassium phosphate (X1), NaCl concentration (X2), and imidazole concentration (X3).

A maximum trimerization ratio (Y1) of approximate A650 nm/A620 nm at 1.024 was obtained at these optimum parameters.

Further examinations, with absorbance spectra, fluorescence spectra and SDS-PAGE, confirmed the presence of bioengineering allophycocyanin trimer with highly trimeric form.

All these results demonstrate that optimized protocol is efficient in purification of bioengineering allophycocyanin trimer with Histag.