A Novel Glucosylation Enzyme: Molecular Cloning, Expression, and Characterization of Trichoderma viride JCM22452 α-Amylase and Enzymatic Synthesis of Some Flavonoid Monoglucosides and Oligoglucos

详细信息    查看全文

• 作者：Akio Noguchi ; Misa Inohara-Ochiai ; Noriko Ishibashi ; Harukazu Fukami ; Toru Nakayama ; Masahiro Nakao
• 刊名：Journal of Agricultural and Food Chemistry
• 出版年：2008
• 出版时间：December 24, 2008
• 年：2008
• 卷：56
• 期：24
• 页码：12016-12024
• 全文大小：330K
• 年卷期：v.56,no.24(December 24, 2008)
• ISSN：1520-5118

文摘

It was found that commercial cellulase preparations from Trichoderma viride showed transglucosylation activity toward (+)-catechin and (−)-epigallocatechin gallate (EGCG) using dextrin as a glucosyl donor. To isolate the enzyme exhibiting transglucosylation activity toward (+)-catechin and EGCG, the present study isolated the cDNA encoding the T. viride JCM22452 α-amylase homologue (TRa2), which showed high amino acid sequence identity to functionally uncharacterized α-amylase homologues from other ascomycetes, which also produced some (+)-catechin and EGCG glucosides. TRa2 was able to glucosylate a wide range of natural flavonoids, particularly (+)-catechin and EGCG, and to hydrolyze maltooligosaccharides (k_cat/K_m for maltotriose, maltotetraose, maltopentaose, maltohexaose, and maltoheptaose were 1.98, 45.2, 58.3, 97.4, and 92.6 s⁻¹ mM⁻¹, respectively) except maltose but could not transfer the monoglucosyl residue to maltooligosaccharides. By ¹H NMR and ¹³C NMR, the structures of several novel glucosides obtained by commercial cellulase preparations from T. viride and TRa2 were determined as (+)-catechin 5-O-α-d-glucopyranoside, (+)-catechin 5-O-α-d-maltoside, (+)-catechin 4′-O-α-d-maltoside, EGCG 5-O-α-d-glucopyranoside, and EGCG 7-O-α-d-maltoside. One of these glucosides, EGCG 5-O-α-d-glucopyranoside, showed higher heat stability and solubility and lower astringency and astringent stimulation than its aglycon, suggesting that EGCG glucosides are functionally superior to EGCG as food additives.