Abstract
Hydrophobins are small proteins (<20 kDa) with an amphipathic tertiary structure that are secreted by various filamentous fungi. Their amphipathic properties provide surfactant-like activity, leading to the formation of robust amphipathic layers at hydrophilic–hydrophobic interfaces, which make them useful for a wide variety of industrial fields spanning protein immobilization to surface functionalization. However, the industrial use of recombinant hydrophobins has been hampered due to low yield from inclusion bodies owing to the complicated process, including an auxiliary refolding step. Herein, we report the soluble expression of a recombinant class I hydrophobin DewA originating from Aspergillus nidulans, and its efficient purification from recombinant Escherichia coli. Soluble expression of the recombinant hydrophobin DewA was achieved by a tagging strategy using a systematically designed expression tag (ramp tag) that was fused to the N-terminus of DewA lacking the innate signal sequence. Highly expressed recombinant hydrophobin DewA in a soluble form was efficiently purified by a modified aqueous two-phase separation technique using isopropyl alcohol. Our approach for expression and purification of the recombinant hydrophobin DewA in E. coli shed light on the industrial production of hydrophobins from prokaryotic hosts.
Highlights
Published: 22 July 2021Hydrophobins are small proteins with an amphipathic structure that were initially isolated from Schizophyllum commune [1]
A method to prepare target gene-specific ramp tags was developed to facilitate the overexpression of several recombinant proteins [26]
The ramp tag comprises a set of rare codons, deduced from codon usage preference in line with copy numbers of tRNA genes in the genome of E. coli K12 and those present in the target gene sequence
Summary
Hydrophobins are small proteins with an amphipathic structure that were initially isolated from Schizophyllum commune [1]. This family of proteins is mainly composed of. Hydrophobins provide a hydrophobic surface layer through self-assembly on the hydrophilic hypha of growing mycelium, when monomers are secreted on the surface of growing mycelia. Hydrophobins are typically divided into two classes (class I and II) depending on hydropathy, solubility, and the structure formed during self-assembly. Both class I and II can form an amphipathic monolayer at a hydrophilic–hydrophobic interface [7,8].
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