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Enzyme Microb Technol. 2014 Jun 10;60:9-15. doi: 10.1016/j.enzmictec.2014.03.009. Epub 2014 Mar 30.

Bioinformatic and biochemical analysis of a novel maltose-forming α-amylase of the GH57 family in the hyperthermophilic archaeon Thermococcus sp. CL1.

Author information

1
Graduate School of Biotechnology and Institute of Life Science and Resources, Kyung Hee University, Yongin 446-701, Republic of Korea.
2
Graduate School of Biotechnology and Institute of Life Science and Resources, Kyung Hee University, Yongin 446-701, Republic of Korea; Research Division for Biotechnology, Korea Atomic Energy Research Institute (KAERI), Jeongeup 580-185, Republic of Korea.
3
Department of Microbiology, University of Massachusetts, Amherst, MA 01003, USA.
4
Graduate School of Biotechnology and Institute of Life Science and Resources, Kyung Hee University, Yongin 446-701, Republic of Korea. Electronic address: cspark@khu.ac.kr.

Abstract

Maltose-forming α-amylase is a glycoside hydrolase family 57 (GH57) member that is unique because it displays dual hydrolysis activity toward α-1,4- and α-1,6-glycosidic linkages and only recognizes maltose. This enzyme was previously identified only in Pyrococcus sp. ST04 (PSMA); however, we recently found two homologs subgroups in Thermococcus species. One subgroup (subgroup A) showed relatively high amino acid sequence similarity to PSMA (>71%), while the other subgroup (subgroup B) showed lower homology with PSMA (<59%). To characterize the subgroup B maltose-forming α-amylase from Thermococcus species (TCMA), we cloned the CL1_0868 gene from Thermococcus sp. CL1 and then successfully expressed the gene in Escherichia coli. Although TCMA has a different oligomeric state relative to PSMA, TCMA showed similar substrate specificity. However, TCMA was shown to hydrolyze maltooligosaccharides more easily than PSMA. Also, TCMA displayed different optimum conditions depending on the glycosidic linkage of the substrate. TCMA had the highest activity at 85°C and at pH 5.0 for α-1,4-glycosidic linkage hydrolysis whereas it showed its maximal activity to cleave α-1,6-glycosidic linkages at 98°C and pH 6.0.

KEYWORDS:

Glycoside hydrolase family 57 (GH57); Hyperthermophile; Maltose-forming α-amylase; Thermococcus sp. CL1

PMID:
24835094
DOI:
10.1016/j.enzmictec.2014.03.009
[Indexed for MEDLINE]

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