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Enzyme Microb Technol. 2018 Jul;114:15-21. doi: 10.1016/j.enzmictec.2018.03.005. Epub 2018 Mar 16.

GH57 amylopullulanase from Desulfurococcus amylolyticus JCM 9188 can make highly branched cyclodextrin via its transglycosylation activity.

Author information

1
Graduate School of Biotechnology and Institute of Life Science and Resources, Kyung Hee University, Yongin, 17140, Republic of Korea.
2
Research Division for Biotechnology, Korea Atomic Energy Research Institute, Jeongeup, 56212, Republic of Korea; Department of Radiation Biotechnology and Applied Radioisotope Science, University of Science and Technology, Daejeon, 34113, Republic of Korea.
3
Gut Microbiome Research Group, Korea Food Research Institute, Sungnam, Republic of Korea.
4
Department of Food and Nutrition, Chungnam National University, Daejon, 34134, Republic of Korea.
5
Department of Pharmaceutical Biology, Institute of Pharmacy and Biochemistry, Johannes Gutenberg University, Mainz, 55128, Germany.
6
Graduate School of Biotechnology and Institute of Life Science and Resources, Kyung Hee University, Yongin, 17140, Republic of Korea. Electronic address: cspark@khu.ac.kr.

Abstract

Desulfurococcus amylolyticus is an anaerobic and hyperthermophilic crenarchaeon that can use various carbohydrates as energy sources. We found a gene encoding a glycoside hydrolase family 57 amylolytic enzymes (DApu) in a putative carbohydrate utilization gene cluster in the genome of D. amylolyticus. This gene has an open reading frame of 1,878 bp and consists of 626 amino acids with a molecular mass of 71 kDa. Recombinant DApu (rDApu) completely hydrolyzed pullulan to maltotriose by attacking α-1,6-glycosidic linkages, and was able to produce glucose and maltose from soluble starch and amylopectin. Although rDApu showed no activity toward α-cyclodextrin (CD) and β-CD, maltooctaose (G8) was detected from reaction with γ-CD. The highest activity of rDApu was measured at pH 5.0 and 95 °C. The half-life of rDApu was 12.7 h at 95 °C and 27 min at 98 °C. Interestingly, rDApu was able to transfer a maltose unit to 6-O-α-maltosyl-β-CD via transglycosylation. Structure analysis using MALDI-TOF/TOF MS and nuclear magnetic resonance revealed that the new transglycosylated products were 61, 64-di-O-maltosyl-β-CD and 61, 63, 65-tri-O-maltosyl-β-CD.

KEYWORDS:

Amylopullulanase; Branched cyclodextrin (CD); Desulfurococcus amylolyticus; Pullulan; Transglycosylation

PMID:
29685348
DOI:
10.1016/j.enzmictec.2018.03.005
[Indexed for MEDLINE]

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