Enzymatic synthesis of ester-linked amino acid-sugar alcohol conjugates for biodegradable polymers

Abstract

Commercial lipases and proteases were tested for the selective acylation of monosaccharides and sugar alcohol through the transesterification of activated amino acid esters. A commercialized protease from Bacillus licheniformis, Optimase M-440 showed the highest catalytic activity in the transesterification of N-t-Boc-L-Phe-OTFE with D-glucose and D-sorbitol. The broad substrate specificity of Optimase M-440 was effective for the acylation of monosaccharides and sugar alcohols with various amino acid esters. Trifluoroethyl ester of L-phenylalanine was more effective for the transesterification than trichloroethyl ester, cyanomethyl ester and oxime ester. Various solvents were tested for the transesterification with Optimase M-440 as a reaction medium. Optimase M-440 catalyzed the highest conversion in pyridine among the tested organic solvents. Optimase M-440 showed a broad substrate specificity towards trifluoroethyl esters of amino acids and the transesterification of L-form of phenylalanine was faster than D-form. Optimase M-440 exhibited a preferable catalytic activity on the primary hydroxyl group of monosaccharides and sugar alcohols and the regioselectivity was confirmed by the $^13C-NMR$ of the conjugates of amino acid and saccharides. The acylation occurred at the primary hydroxyl groups of D-glucose, and regioselectivity and chemoselectivity of Optimase M-440 was observed in acylation of D-glucosamine. Optimase M-440 catalyzed the acylation of two primary hydroxyl groups of sugar alcohols in spite of the change of reaction conditions. Sugar alcohol conjugates of N-t-Boc-L-Phe-OTFE and N-t-Boc-L-Met-OTFE were synthesized without the acylation of secondary hydroxyl groups. In the transesterification of Nα, Nε-di-t-Boc-L-Lys-OTFE$ and N-t-Boc-L-Asp-diOTFE, Optimase M-440 did not discriminate the glycosidic hydroxyl groups. Only when the glycosidic -OH was blocked, Optimase M-440 could selectively catalyze the acylation of primary hydroxyl group without o...