Ｐｅｎｉｃｉｌｌｉｕｍ ｃｉｔｒｉｎｕｍが生産するｅｎｄｏ‐１，４‐β‐Ｄ‐ｇａｌａｃｔａｎａｓｅの糖転移反応

Abstract

1. Two galactanases (I and II) from Penicillium citrinum were purified to a homogeneous state. These enzymes were electrophoretically distinctive, but were very similar in proteochemical and enzymatic properties. The enzymes hydrolyzed soybean arabinogalactan (SAG) in an endo manner. They also acted on o-nitrophenyl-β-D-galactoside (ONPG), p-nitrophenyl-β-Dgalactoside and, β-1, 4-galactobiose (Gal2) after lag phases. 2. The action of galactanase I on ONPG was studied. In the course of the reaction, the accumulation of various transfer products was observed by thin-layer chromatography. The products were isolated from the reaction mixture and their structures were examined. They were a series of β-1, 4-linked galactooligosaccharides (Gal2-Gal4) and o-nitrophenol (ONP)-substituted oligosaccharides (G2-ONP-G5-ONP). The enzyme liberated ONP without significant lag phases from G2-ONP-G5-ONP at a much faster rate than that from ONPG. Furthermore, the lag phase of ONP liberation from ONPG could be eliminated by the addition of Gala or Gal4 to the mixture. Thus a reaction mechanism that involved transfer reaction in addition to hydrolysis was proposed for the cause of the observed lag . 3. The transfer reaction catalyzed by galactanase I was studied using SAG as a donor. The enzyme had a broad acceptor specificity and gave transfer products from several alcohols, monosaccharides, sugar alcohols, glycerol and its derivatives, and catechol. Formation of transfer products from SAG and glycerol was examined in detail. In the course of the reaction, transfer products with various degrees of polymerization were accumulated, suggesting the enzyme activity as transferring galactosyl and galactooligosyl units to the acceptor. The amounts of transfer products depended on the glycerol concentration, and about 50% of the galactose residues which could be liberated from SAG by the enzyme were transferred to glycerol at an acceptor concentration of 2.5%. Two transfer products were isolated and their structures were examined. They were 2-O-β-galactosyl glycerol and 2-O-β-galactobiosyl glycerol, whereas the transfer product from ONPG and glycerol by Escherichia coli β-galactosidase was 1-O-β-galactosyl glycerol.