Increased solubility of integrin betaA domain using maltose-binding protein as a fusion tag.
In proteomics research, generation of recombinant proteins in their native, soluble form with large quantity is often a challenging task. To tackle the expression difficulties, different expression vectors with distinct affinity fusion tags, i.e. pET-43.1a (N-utilization substance A tag), pMAL-cRI (maltose binding protein tag) (MBP tag), pGEX-4T-2 (glutathione S-transferase tag), and pET-15b (hexahistidine tag) were compared for their effects on the productivity and solubility, which were assessed by SDS-PAGE and immunoblotting, of the integrin betaA domain. The incubation temperatures were tested for its effects on these parameters. Our data suggested that MBP tag enhanced the yield and solubility of the betaA domain protein, which can also be recognized using an anti-CD18 antibody, at room temperature incubation. Thus, the nature of fusion partner chosen for expression in bacteria and its incubation temperature would significantly affect the yield and solubility of the recombinant target protein.
Lee
Nikki P Y
NP
Department of Surgery, The University of Hong Kong, Pokfulam, Hong Kong.
Tsang
Stella
S
Cheng
R Holland
RH
Luk
John M
JM
D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002846:Chromatography, Affinity; D006801:Humans; D039641:Integrin beta Chains / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D007962:Leukocytes / Q000166:cytology / Q000378:metabolism; D058619:Maltose-Binding Proteins; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D012995:Solubility
D002352:Carrier Proteins; D039641:Integrin beta Chains; D058619:Maltose-Binding Proteins; D011993:Recombinant Fusion Proteins