Monogalactosyldiacylglycerol (MGDG) synthase (UDPgalactose:1,2-diacylglycerol 3--D-galactosyltransferase; EC 2.4.1.46) catalyzes formation of MGDG, a major structural lipid of chloroplast. We cloned a cDNA for the synthase from cucumber cDNA library. The full-length cDNA clone was 2142 bp, and it contains a 1575-bp open reading frame encoding 525 aa. The open reading frame consists of the regions for a mature protein (422 aa; M r of 46,552) and transit peptide to chloroplast (103 aa). Although the molecular weight of mature protein region matched that purified from cucumber cotyledons, it was quite different from those purified from spinach (Ϸ20 kDa) reported by other groups. The mature region of the protein was expressed in Escherichia coli as a fusion protein with glutathione Stransferase. The expression in E. coli showed that the protein catalyzed MGDG synthesis very efficiently. Therefore, we concluded that the cDNA encodes MGDG synthase in cucumber. In addition, the deduced amino acid sequence of the MGDG synthase cDNA showed homology with MurG of Bacillus subtilis and E. coli, which encode a glycosyltransferase catalyzing the last step of peptidoglycan synthesis in bacteria. This sequence homology implies that the machinery of chloroplast membrane biosynthesis is evolutionarily derived from that of cell wall biosynthesis in bacteria. This is consistent with the endosymbiotic hypothesis of chloroplast formation.Chloroplast thylakoid membranes are the site of the plant's photochemical reactions and are composed of proteins, such as the photosynthetic reaction center, and lipids. In higher plants and eukaryotic algae, about 50% (wt͞wt) of the membrane lipids of chloroplasts are two major galactolipids, monogalactosyldiacylglycerol (MGDG), and digalactosyldiacylglycerol (DGDG). MGDG and DGDG are not only major constituents of chloroplast membrane but have a major role in determining the physicochemical characteristics of thylakoid membranes (1, 2). A tightly bound MGDG molecule in the photosystem II reaction center complex has been identified as important in photosynthesis (3).The final step of MGDG synthesis is catalyzed by MGDG synthase (UDPgalactose: 1,2-diacylglycerol 3--D-galactosyltransferase; EC 2.4.1.46). The enzyme transfers a galactose from UDPgalactose to 1,2-diacylglycerol in chloroplast envelope (4). Another major galactolipid, DGDG, is synthesized by dismutation of two molecules of MGDG (5). We have found that MGDG synthase activity in cucumber seedlings dramatically increased coincidentally with the accumulation of the two major galactolipids, MGDG and DGDG, during chloroplast development (6). This fact and other reports (4) strongly suggest that the reaction catalyzed by MGDG synthase is a key step in the formation of chloroplast membrane. Despite the importance of MGDG synthase for chloroplast membrane biogenesis, molecular characteristics of the enzyme remain ambiguous. Purification of MGDG synthase from spinach envelopes was reported by two groups (7,8). According to their results,...
The separation and identification of pigments, chlorophylls, and carotenoids of seven teas and fresh leaf of tea (Camellia sinensis) by high-performance liquid chromatography (HPLC) are described. HPLC was carried out using a Symmetry C(8) column with a photodiode array detector. Pigments were eluted with a binary gradient of aqueous pyridine solution at a flow rate of 1.0 mL/min at 25 degrees C. HPLC analyses achieved the separation of more than 100 pigment peaks, and 79 pigment species, 41 chlorophylls, and 38 carotenoids were detected. The presence of degraded chlorophylls was a common feature, and the number and the variety of pigments differed with tea species. Generally, the numbers of chlorophyll species tended to increase with processing steps, while carotenoid species were decreased, especially by heating. Particularly in green teas, a change of carotenoid structure, conversion of violaxanthin to auroxanthin, occurred. In hot water extracts of teas, both chlorophylls and carotenoids were also detected, but the concentration of chlorophylls was less than 2% as compared with acetone extracts. The pigment compositions were compared between tea species, and they are discussed in terms of the differences in their manufacturing processes.
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