Metal compound-mediated hydrolytic cleavage of oxidized insulin B chain: Regioselectivity and influence of peptide secondary structure

The interaction of oxidized insulin B chain (B) with cis-[Pd(en)Cl-2] (en = ethylenediamine), cis-[Pd-(dtco-3-OH)Cl-2] (dtco-3-OH = dithiacyclooctan-3 -ol) and CuCl2 was studied by electrospray mass spectrometry. It is discove red that the binding of Pd(II) complexes and the sites of cleavage are high ly dependent on the secondary structure and local environment of B. The hyd rolytic cleavage of denatured B by Pd(II) complexes was monitored by HPLC. The reaction is regioselective and follows first order kinetics with half-l ife of 4.8 days at 40 degreesC. Two amide bonds, i.e. at Leu6-Cys7 and at G ly8-Ser9, which are close to the two potential Pd(II) binding sites His5 an d His10, are selectively cleaved. In the case of Cu(II) ion as promoter, on ly one cleavage site was observed which is located at Gly8-Ser9 bond. These results provide improved understanding on the design of artificial metallo peptidase.