Fmoc-D-Orn(Boc)-Alko-PEG Resin

In the movement to decarbonize our economy and move away from fossil fuels we will need to harness the waste products of our activities, such as waste lignocellulose, methane, and carbon dioxide. Our wastes need to be integrated into a circular economy where used products are recycled into a manufacturing carbon cycle. Key to this will be the recycling of plastics at the resin and monomer levels. Biotechnology is well suited to a future chemical industry that must adapt to widely distributed and diverse biological chemical feedstocks. Our increasing mastery of biotechnology is allowing us to develop enzymes and organisms that can synthesize a widening selection of desirable bulk chemicals, including plastics, at commercially viable productivities. Integration of bioreactors with electrochemical systems will permit new production opportunities with enhanced productivities and the advantage of using a low-carbon electricity from renewable and sustainable sources.

Recently, an unexpected modified residue, gamma-hydroxy-D-valine (D-Hyv), was identified within ribosomally expressed polypeptide chains of four conopeptides from the venoms of Conus gladiator and Conus mus. To assemble Hyv-containing peptides, we have explored several routes for the synthesis of appropriately functionalized Hyv building blocks. D-Hyv was produced from D-Val by using a variation of the previously published K2PtCl4/CuCl2 oxidative method. Direct synthesis of Boc- or Cbz-D-Hyv lactone proceeded in low yield; additionally, the lactones are too unreactive for solid-phase applications. 9-Borabicyclononane or copper-complexed D-Hyv was prepared and treated with tert-butyldimethylsilyl trifluoromethanesulfonate (TBDMSOTf) to produce D-Hyv(O-TBDMS). The most efficient complex disruption was achieved by Chelex 110 resin (Na+ form) treatment of copper-complexed D-Hyv(O-TBDMS). Reaction of D-Hyv(O-TBDMS) with Fmoc-OSu produced Fmoc-D-Hyv(O-TBDMS) in 26% yield from D-Val. The Fmoc-D-Hyv(O-TBDMS) diastereomers were separated by preparative RP-HPLC in 13% yield from D-Val. Fmoc-D-Hyv(O-TBDMS) was used for the synthesis of the conopeptide gld-V* from Conus gladiator. The isolated synthetic and natural products had coincidental mass and NMR spectra. The methodology presented herein will greatly facilitate biological studies of Hyv-containing sequences, such as receptor responses to hydroxylated versus nonhydroxylated conopeptides and the relative susceptibility of proteins to modification by oxidative stress.

Aldoses and ketoses can glycate proteins yielding isomeric Amadori and Heyns products, respectively. Evidently, D-fructose is more involved in glycoxidation than D-glucose favoring the formation of advanced glycation endproducts (AGEs). While Amadori products and glucation have been studied extensively, the in vivo effects of fructation are largely unknown. The characterization of isomeric Amadori and Heyns peptides requires sufficient quantities of pure peptides. Thus, the glycated building block Nα-Fmoc-Lys[Nε-(2-deoxy-D-glucos-2-yl),Nε-Boc]-OH (Fmoc-Lys(Glc,Boc)-OH), which was synthesized in two steps starting from unprotected D-fructose and Fmoc-L-lysine hydrochloride, was site-specifically incorporated during solid-phase peptide synthesis. The building block allowed the synthesis of a peptide identified in tryptic digests of human serum albumin containing the reported glycation site at Lys233. The structure of the glycated amino acid derivatives and the peptide was confirmed by mass spectrometry and NMR spectroscopy. Importantly, the unprotected sugar moiety showed neither notable epimerization nor undesired side reactions during peptide elongation, allowing the incorporation of epimerically pure glucosyllysine. Upon acidic treatment, the building block as well as the resin-bound peptide formed one major byproduct due to incomplete Boc-deprotection, which was well separated by reversed-phase chromatography. Expectedly, the tandem mass spectra of the fructated amino acid and peptide were dominated by signals indicating neutral losses of 18, 36, 54, 84 and 96 m/z-units generating pyrylium and furylium ions.