Selective hydrolysis of oxidized insulin chain B by a Zr(iv)-substituted Wells–Dawson polyoxometalate

Abstract

We report for the first time on the selective hydrolysis of a polypeptide system by a metal-substituted polyoxometalate (POM). Oxidized insulin chain B, a 30 amino acid polypeptide, was selectively cleaved by the Zr(IV)-substituted Wells-Dawson POM, K15H[Zr(α2-P2W17O61)2]·25H2O, under physiological pH and temperature conditions in aqueous solution. HPLC-ESI-MS, LC-MS/MS, MALDI-TOF and MALDI-TOF MS/MS data indicate hydrolysis at the Phe1-Val2, Gln4-His5, Leu6-Cys(SO3H)7, and Gly8-Ser9 peptide bonds. The rate of oxidized insulin chain B hydrolysis (0.45 h(-1) at pH 7.0 and 60 °C) was calculated by fitting the integration values of its HPLC-UV signal to a first-order exponential decay function. (1)H NMR measurements show significant line broadening and shifting of the polypeptide resonances upon addition of the Zr(IV)-POM, indicating that interaction between the Zr(IV)-POM and the polypeptide takes place in solution. Circular dichroism (CD) measurements clearly prove that the flexible unfolded nature of the polypeptide was retained in the presence of the Zr(IV)-POM. The thermal stability of the Zr(IV)-POM in the presence of the polypeptide chain during the hydrolytic reaction was confirmed by (31)P NMR spectroscopy. Despite the highly negative charge of the Zr(IV)-POM, the mechanism of interaction appears to be dominated by a strong metal-directed binding between the positively charged Zr(IV) center and negatively charged amino acid side chains.

Selectivity and Reactivity of ZrIV and CeIV Substituted Keggin Type Polyoxometalates Toward Cytochrome c in Surfactant Solutions

“…Ce IV and Zr IV substituted POMs were shown to promote selective hydrolysis of a variety of water soluble proteins, ranging from flexible polypeptide such as oxidized insulin chain B (Sap et al, 2015), to larger, globular proteins such as hen egg white lysozyme (Stroobants et al, 2013, 2014d), human serum albumin (HSA) (Goovaerts et al, 2013; Stroobants et al, 2014a,b), horse heart cytochrome c (Cyt c) (Sap et al, 2016) and horse heart myoglobin (Ly et al, 2015a; Ly and Parac-Vogt, 2017). However, the potential of these POMs to act as artificial proteases toward hydrolysis of proteins in the presence of surfactants is still largely unexplored.…”

Metal assisted peptide bond hydrolysis: Chemistry, biotechnology and toxicological implications

“…Oxidized insulin chain B was chosen as a next model for hydrolysis by Zr(IV)-substituted Wells-Dawson POM [81]. Four cleavage sites were observed for reaction performed at 37 • C (Phe1-Val2, Gln4-His5, Leu6-Cys(SO 3 H)7, Gly8-Ser9) and five at 60 • C (additionally Gly20-Glu21).…”

Comparative Study of the Reactivity of Zirconium(IV)‐Substituted Polyoxometalates towards the Hydrolysis of Oligopeptides

“…The demonstrated trend in selectivity can be best compared to that observed for oxidized insulin chain B 12. The selectivity of hydrolysis was also driven by coordination chemistry between the Zr IV ion in the POM and the polypeptide, whereas no evidence of specific electrostatic interactions, as is the case for protein hydrolysis, was observed.…”

“…The sandwich 1:2 Zr IV ‐substituted Wells–Dawson POM was the first POM that exhibited peptide bond hydrolysis in a variety of dipeptides and oligopeptides under mildly acidic conditions 8b,8f,8g,12. Their hydrolysis was significantly accelerated owing to the binding of the Zr IV ion to the amine nitrogen atom and the amide carbonyl oxygen atom.…”