A family 1 galactosyl hydrolase performs galactosyltransferase activity requiring Mg2+ and conferring freezing tolerance (617.8)

Abstract

The two most abundant photosynthetic thylakoid membrane lipids are monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG). Understanding galactolipid metabolism is therefore critical to understanding thylakoid formation and subsequent photosynthetic activity. The first enzymatic activity described to synthesize DGDG was named galactolipid: galactolipid galactosyl transferase (GGGT), as it removed a galactosyl head group from MGDG and transferred it to another MGDG making DGDG. Though it is now understood that a separate enzyme, DGD1, synthesizes most of the DGDG in vivo, GGGT was re‐identified in Arabidopsis as the gene product of SENSITIVE TO FREEZING 2 (SFR2). SFR2 was shown to protect Arabidopsis from freezing damage, to be located on the chloroplast outer envelope and to be similar to family‐1 β‐glycosidases. Here, we present a model of the 3‐dimensional structure of SFR2 based on its homology to family‐1 β‐glycosidases and show the functional significance of several unique regions. Using the model, we identify residues required for SFR2 activity that are not conserved among family‐1 β‐glycosidases. We experimentally define pH and temperature optimums for enzymatic activity in vitro. Unexpectedly, we show that SFR2 is a metalloenzyme, unlike its family members. Our data suggest that while SFR2 retains the core structure and active site residues of a family‐1 β‐glycosidase, it has unique residues, regions, and co‐factors required to alter its function to that of a transferase.Grant Funding Source: Supported by the Department of Energy