Peptide Synthesis in Water IV. Preparation of N-Ethanesulfonylethoxycarbonyl (Esc) Amino Acids and Their Application to Solid Phase Peptide Synthesis

Abstract

Synthesis of peptides was an arduous task before development of the solid phase method. The solid phase method has facilitated automated peptide synthesis and the development of combinatorial chemistry. It has many advantages compared with classical solution peptide synthesis, but requires a large amount of organic solvents. Disposal of organic solvents is one of many important environmental problems, therefore we decided to perform peptide synthesis in water. To perform synthesis in water, protected amino acids must be water-soluble. We have studied water-soluble N-protecting groups and we developed 2-[phenyl(methyl)sulfonio]ethoxycarbonyl (Pms) 1,2) as a water-soluble N-protecting group. Since the Pms group is an onium salt, it is rather unstable in comparison with N-protecting groups which are in general use (such as t-butoxycarbonyl group, 3) benzyloxycarbonyl group 4) and 9-fluorenylmethoxycarbonyl (Fmoc) group 5) ) for peptide synthesis. Here we report preparation of the ethanesufonylethoxycarbonyl (Esc) group and its application to peptide synthesis in water.Tesser and Balvert-Geers 6) reported the methanesulfonylethoxycarbonyl (Msc) group as a readily removable Nprotecting group by treatment with 0.1 or 0.2 N NaOH solution. This protecting group is hydrophilic rather than hydrophobic. We designed the Esc group (Fig. 1), expecting it to display both hydrophilic and hydrophobic character (i.e. we expect Esc-amino acids to exhibit reasonable solubility in aqueous and organic solvents). The Esc group was introduced onto amino acids by the reaction of 2-ethanesulfonylethyl chloroformate (Esc-Cl) as shown in Chart 1 (Route 1). 2-Ethanesulfonylethanol was converted to Esc-Cl by treatment with phosgene (prepared from triphosgene) 7) in dichloromethane. Since Esc-Cl was labile and decomposed during purification, it was used without purification for the next acylating reaction. Esc-Cl, H-Phe-OH and triethylamine were allowed to react in aqueous 50% acetonitrile to give Esc-Phe-OH. HPLC analysis of the crude product exhibited impurities and the product could not be purified by silica gel column chromatography and recrystallization. Pure Esc-Phe-OH was obtained by RP-HPLC purification and the yield was 38%. To improve the reaction yield and purification procedure, another synthetic route (Route 2 in Chart 1) to the Escamino acid was studied. 2-Ethanesulfonylethanol was reacted with 4-nitrophenyl chloroformate to give ethanesulfonylethyl-4-nitrophenyl carbonate (Esc-ONp). The crude EscONp was stable and could be purified by silica gel column chromatography or recrystallization from a mixture of ether and hexane. The yield was 78%. Other Esc-amino acids (Esc-Leu-OH, Esc-Gly-OH, Esc-Tyr-OH) were also prepared using Esc-ONp and their analytical data are shown in Table 1. Esc-Gly-OH and Esc-Tyr-OH were soluble in water and Esc-Leu-OH and Esc-Phe-OH were not readily soluble in water. Esc-Leu-OH and Esc-Phe-OH were soluble in aqueous 2% Triton X. To evaluate Esc-amino acids, Leu-enkephalin amide was synthesized by the sol...

Flow‐based SPPS for protein synthesis: A perspective

“…[ 93 ] The first efforts on water‐based SPPS were reported by Kawasaki and co‐workers, who developed water‐compatible coupling agents and protecting groups. [ 108–115 ] Notably, PEG‐based CM‐resin, which is often used in flow‐based SPPS, and several related resins show good swelling properties in water. To further improve the properties of water‐compatible protecting groups, SO 3 − ‐functionalized protecting groups, so‐called 9‐(2‐sulfo)fluorenylmethyloxycarbonyl (Smoc) amino acid derivatives, [ 116 ] as well as additional solvation strategies, additional compatible activators and microwave irradiation were successfully implemented.…”

Greener solvents for solid-phase organic synthesis

“…1). The 2-ethanesulfonylethoxycarbonyl (Esc) 2 group was found to be moderately base labile and moderately polar, giving it good solubility in aqueous and organic solutions, producing the test peptide Leu-enkephalinamide in a 71 % isolated yield [19]. However, the lack of aromatic functionality, excluding the aromatic amino acids, made it undesirable at the time as they are not detectable by optical absorbance spectroscopy, making them unsuitable for automated SPPS.…”

Peptide synthesis ‘in water’ by a solution‐phase method using water‐dispersible nanoparticle Boc‐amino acid

“…In conventional organic syntheses, reactant molecules are routinely dissolved in solvents to carry out the reaction efficiently. Peptide synthesis can be carried out in water via chemical conversion of protected amino acids to water‐soluble forms4–7. This procedure, however, requires an additional conversion step in the synthetic process, and this is undesirable in terms of preparation cost, resource saving and energy conservation.…”