Intein-mediated rapid purification of recombinant maxadilan and M65 and their acute effects on plasma glucose

Maxadilan is a potent vasodilatory peptide present in the salivary glands of the sand fly. Maxadilan and M65, a deletion variation of maxadilan, are agonist- and antagonist-specific for the PAC1 receptor. In order to obtain the recombinant maxadilan and M65 efficiently by intein-mediated single column purification, the genes encoding maxadilan and M65 were designed, synthesized and cloned into Escherichia coli expression vector pKYB. The recombinant maxadilan and M65 with homogeneity over 95% were released from the chitin-bound intein tag by β-mercaptoethanol. Intraperitoneal injection of the recombinant maxadilan caused an acute elevation of plasma glucose, imitating pituitary adenylate cyclase-activating polypeptide (PACAP) 27, in NIH mice, while the VPAC1-agonist and VPAC2-agonist had no significant effects on the levels of plasma glucose. M65 alone had no effect on the plasma glucose, but blocked the glucose excursion caused by maxadilan by 12.7% and blocked the glucose excursion caused by the PACAP 27 by 11.6%. The acute effects of the recombinant maxadilan and M65 on the plasma glucose indicated that they had the characteristics as the agonist and antagonist for PAC1.