Comparative Functional Characterization of LuUGT74S1 from Linum usitatissimum L. and Its Closed Homologues from Fragaria vesca Subsp. vesca and Camellia sinensis

Abstract

The bioactive lignan secoisolariciresinol-diglucoside (SDG) accumulates in flaxseed (Linum usitatissimum), where LuUGT74S1 is involved in its formation by the sequential transfer of two glucose units to the aglycone secoisolariciresinol (SECO). Here, we investigated whether similar proteins catalyze the identical reaction in other SDG-producing plants. A phylogenetic analysis identified FvUGT74DH1 from Fragaria vesca subsp. vesca and CsUGT74DG1 from Camellia sinensis as LuUGT74S1-like. Recombinant LuUGT74S1 glucosylated (−)- and (+)-SECO to SDG, but none of the seven selected substrate-analogues. The LuUGT74S1-homologous proteins failed to glucosylate SECO but were active toward substrates structurally related to SECO. In contrast to CsUGT74DG1 and FvUGT74DH1, which were active in Escherichia coli, LuUGT74S1 was unable to convert SECO in intact transgenic E. coli cells, presumably due to the inhibitory Mg²⁺ concentration in the bacteria. LuUGT74S1 has become highly substrate-specific, probably due to selection pressure, while its homologues remained promiscuous. The SECO-glucosylating enzymes must have evolved through convergent evolution in different species.