Microwave-assisted solid-phase peptide synthesis of neurosecretory protein GL composed of 80 amino acid residues

Abstract

We recently identified a novel cDNA encoding a small secretory protein of 80 amino acid residues, termed neurosecretory protein GL (NPGL), from the chicken hypothalamus. Homologs of NPGL have been reported to be present in mammals, such as human and rat. NPGL is amidated at its C-terminus, contains an intramolecular disulfide bond, and is hydrophobic in nature. In this study, we have optimized the synthesis of the entire 80-amino acid peptide sequence of rat NPGL by microwave-assisted solid-phase peptide synthesis. NPGL was obtained with a 10% yield when the coupling reactions were performed using 1-[Bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium-3-oxid hexafluorophosphate (HATU) at 50 °C for 5 min, and Fmoc deprotections were performed using 40% piperidine containing 0.1 M HOBt. Furthermore, the disulfide bond of NPGL was formed with 20% yield with the use of glutathione-containing redox buffer and 50% acetonitrile.

Neurosecretory protein GL stimulates food intake, de novo lipogenesis, and onset of obesity

“…Rat NPGL of 80 amino acid residues, its analogs NPGL-Gly and NPGL(32-80), and NPGM ( Figure 1—figure supplement 1E ) were synthesized by microwave-assisted solid-phase peptide synthesis using an automated peptide synthesizer (Syro Wave; Biotage, Uppsala, Sweden) as previously described ( Masuda et al, 2015 ).…”

Production and characterization of neurosecretory protein GM usingEscherichia coliand Chinese Hamster Ovary cells

“…The specificity of the antibody was confirmed by dot-immunoblot analysis. Rat NPGL [6] and NPGM were blotted on a nitrocellulose membrane and probed with the antibody against NPGM (1:250 dilution) and revealed with an alkaline phosphatase-conjugated secondary antibody (1:1000 dilution).…”

Synthesis and Bioactivity of Diastereomers of the Virulence Lanthipeptide Cytolysin

“…Hydrophobic peptides are prone to incomplete coupling during solid phase peptide synthesis (SPPS) because of inaccessibility of the reagents to the N-terminus of the elongating peptide chain. 9 Secondly, the structures of cytolysins contain a dehydro amino acid in the second position within the thioether rings. Dehydro amino acids cannot be incorporated via the usual elongation methods of Fmoc SPPS because the enamine liberated upon Fmoc deprotection is very unreactive and would tautomerize to the imine followed by hydrolysis to the ketone, preventing further peptide coupling.…”