Isotopic Combinatomer Analysis Provides in Vivo Evidence of the Direct Epimerization of Monoglucosyl Diacylglycerol in Cyanobacteria.

Abstract

Galactolipids constitute the majority of photosynthetic membranes called thylakoid membranes in cyanobacteria and chloroplasts of land plants and algae. The galactolipids, although identical in headgroup structure, are synthesized by significantly different pathways in cyanobacteria and chloroplasts. In the cyanobacterial pathway, monoglucosyl diacylglycerol (GlcDG) is synthesized first and then converted to monogalactosyl diacylglycerol (MGDG). On the basis of circumstantial evidence, the mechanism of conversion was thought to be epimerization at C-4, but no direct evidence has yet been provided, because there is no in vitro enzymatic system of the putative membrane-bound reaction. Labeling studies with 14C and 13C suggested that the labels in the headgroup and the acyl groups were kept at a reasonably constant ratio before and after the conversion. We then provide in vivo evidence of the direct epimerization based on detailed isotopomer analysis of the conversion, named "combinatomer analysis". The different types of molecules formed by the combination of labeled or unlabeled parts (sn-1 acyl, sn-2 acyl, glycerol, and hexose) are called here "combinatomers". Combinatomer analysis of the experiments with pulse labeling with 13C and chase in Anabaena sp. PCC 7118 indicated that the composition of combinatomers in the precursor GlcDG was kept unchanged in the product MGDG. Production of combinatomers resulting from exchange of hexose was minimal. This provides solid evidence of the epimerization of the glucose moiety of GlcDG, as well as the direct desaturation of acyl groups at the sn-1 position.