Chemical synthesis of per‐selenocysteine human epidermal growth factor

Abstract

Human seleno‐epidermal growth factor (seleno‐EGF), a 53‐residue peptide where all six cysteine residues of the parent human EGF sequence were replaced by selenocysteines, was synthesized and the oxidative folding of a polypeptide containing three diselenide bonds was compared to that of the parent cysteine peptide. The crude high performance liquid chromatography (HPLC) profiles clearly showed that both the native EGF and its selenocysteine‐analogue fold smoothly, yielding a single sharp peak, proving that even in the case of three disulfide‐bonded polypeptides the disulfide‐to‐diselenide bond substitution is highly isomorphous, as confirmed by conformational circular dichroism measurements and particularly by the biological assays.

Unexpected diselenide metathesis in selenocysteine-substituted biologically active peptides

“…36 We previously reported the synthesis of Sec-substituted epidermal growth factor (seleno-EGF) in which all three disulfide bonds of the native EGF were substituted with diselenide bonds. 24 Like seleno-apamin, no isomer was observed during oxidative folding, showing the high stability of the native tertiary structure of seleno-EGF. It seems that in the case of seleno-peptides with stable tertiary structure, diselenide metathesis is not observed.…”

“…Other diselenide-substituted biologically active peptides and proteins have been synthesized, which demonstrated the utility of the diselenide-substitution strategy to stabilize the structure of naturally-occurring peptides and proteins. [19][20][21][22][23][24] Conversely, the diselenide bond possesses a lower bond energy compared with the disulfide bond (diselenide bond: 172 kJ mol −1 , disulfide bond: 240 kJ mol −1 ). 25 Xu et al showed the visible light-induced diselenide metathesis reaction to demonstrate the utility of diselenide compounds for dynamic combinatorial chemistry (DCC) and reported that the reaction proceeds by a radical mechanism.…”

“…Other diselenide-substituted biologically active peptides and proteins have been synthesized, which demonstrated the utility of the diselenide-substitution strategy to stabilize the structure of naturally-occurring peptides and proteins. 19–24…”

Selenium in Peptide Chemistry

“…Walewska et al [ 67 ] demonstrated that pairwise replacement of Cys with Sec in µ-conotoxin SIIIA enhanced the accumulation of natively folded proteins and decreased the number of intermediates. The substitution of the diselenide crosslinker for the interchain disulfide bridge (A6–A11) in human insulin was reported to ameliorate the challenge of efficiently combining the A and B chains to yield native insulin in a relatively short time and with a higher yield, which was accompanied by increased stability of the protein compared to the wild-type analog [ 68 ], as also shown in caenopore-5 (Cp-5) [ 69 ], human epidermal growth factor (EGF) [ 70 ], and α-conotoxin [ 71 ] studies. Interchain substitution on a single chain is indeed a more straightforward strategy than the synthetic method developed by Arai et al [ 72 ], who modified both A and B chains ([C7U A ,C7U B ]) of bovine pancreatic insulin (BPIns).…”