Design, Preparation Of Monomeric Insulin B27 Lys DTrI & Dopamine Transporter Antagonist F(Phenyl)YTPT(Benzyl)G

Monomeric B27 Lys destripeptide insulin (B27 Lys DTrI) was designed in order to being produced from its single chain precursor through tryptic hydrolysis instead of the less efficient tryptic transpeptidation. Semisynthetic B27 Lys DTrI exhibited superior monomeric property in size-exclusion chromatography, and its in vivo biological activity was 80% of insulin determined by mouse convulsion assay. The monomeric B27 Lys DTrI precursor (MIP) expressed in Pichia pastoris was purified from the cultured medium, and was converted to B27 Lys DTrI by tryptic hydrolysis. The yield of MIP was 200 mg per liter of culture, and the overall yield of purified B27 Lys DTrI from the crude MIP was 70%. The monomeric property and in vivo biological activity of recombinant B27 Lys DTrI were identical to that obtained by semisynthesis. F(Phenyl)YTPT(Benzyl)G is found to be a new dopamine transporter antagonist. Dopamine transporter was shown to play an important role in the pathophysiology of mood disorders and drug abuse, including depression and cocaine dependent. It has been suggested that both agonists and antagonists of dopamine transporter have strong therapeutic potential, but the de novo design rules of these regulators are unclear. Here we report the design of F(Phenyl)YTPT(Benzyl)G, a selective and potent peptidic inhibitor of dopamine transporter generated via aromatic modification of insulin B (24-30) peptide scaffold. Our results demonstrated that the aromatic modification can constrain the peptide into β-turn conformation, which is biased by dopamine transporter. The lipophilic peptide can penetrate the blood-brain barrier in mice to block...