Extra terminal residues have a profound effect on the folding and solubility of a Plasmodium falciparum sexual stage-specific protein over-expressed in Escherichia coli.

The presence of extra N- and C- terminal residues can play a major role in the stability, solubility and yield of recombinant proteins. Pfg27 is a 27K soluble protein that is essential for sexual development in Plasmodium falciparum. It was over-expressed using the pMAL-p2 vector as a fusion protein with the maltose binding protein. Six different constructs were made and each of the fusion proteins were expressed and purified. Our results show that the fusion proteins were labile and only partially soluble in five of the constructs resulting in very poor yields. Intriguingly, in the sixth construct, the yield of soluble fusion protein with an extended carboxyl terminus of 17 residues was several fold higher. Various constructs with either N-terminal or smaller C-terminal extensions failed to produce any soluble fusion protein. Furthermore, all five constructs produced Pfg27 that precipitated after protease cleavage from its fusion partner. The sixth construct, which produced soluble protein in high yields, also gave highly stable and soluble Pfg27 after cleavage of the fusion. These results indicate that extra amino acid residues at the termini of over-expressed proteins can have a significant effect on the folding of proteins expressed in E. coli. Our data suggest the potential for development of a novel methodology, which will entail construction of fusion proteins with maltose binding protein as a chaperone on the N-terminus and a C-terminal 'solubilization tag'. This system may allow large-scale production of those proteins that have a tendency to misfold during expression.