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Purification and molecular characterization of cold-active β-galactosidase from Arthrobacter psychrolactophilus strain F2

Abstract

In this study, we purified and molecularly characterized a cold-active β-galactosidase from Arthrobacter psychrolactophilus strain F2. The purified β-galactosidase from strain F2 exhibited high activity at 0°C, and its optimum temperature and pH were 10°C and 8.0, respectively. It was possible to inactivate the β-galactosidase rapidly at 45°C in 5 min. The enzyme was able to hydrolyze lactose as a substrate, as well as o-nitrophenyl-β-d-galactopyranoside (ONPG), the K m values with ONPG and lactose being calculated to be 2.8 mM and 50 mM, respectively, at 10°C. Moreover, the bglA gene encoding the β-galactosidase of strain F2 was cloned and analyzed. The bglA gene consists of a 3,084-bp open reading frame corresponding to a protein of 1,028 amino acid residues. BglAp, the gene product derived from bglA, had several conserved regions for glycosyl hydrolase family 2, e.g., the glycosyl hydrolase 2 (GH2) sugar binding domain, GH2 acid-base catalyst, GH2 triosephosphate isomerase barrel domain, GH2 signature 1, and several other GH2 conserved regions. From these facts, we conclude that the β-galactosidase from A. psychrolactophilus strain F2, which is a new member of glycosyl hydrolase family 2, is a cold-active enzyme that is extremely heat labile and could have advantageous applications in the food industry.

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Acknowledgements

We are indebted to Professor Toshihiro Watanabe, Tokyo University of Agriculture, for his help in the amino acid sequence determination. We are also grateful to Ms. Kumiko Sato, Ms. Takae Tamura, Ms. Riwako Shima, and Mr. Takahito Hayashi for their skillful assistance. This research was supported in part by a grant from the Northern Advancement Center for Science & Technology to T.N.

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Correspondence to Tomoyuki Nakagawa.

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Nakagawa, T., Fujimoto, Y., Ikehata, R. et al. Purification and molecular characterization of cold-active β-galactosidase from Arthrobacter psychrolactophilus strain F2. Appl Microbiol Biotechnol 72, 720–725 (2006). https://doi.org/10.1007/s00253-006-0339-0

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Keywords

  • Lactose
  • Arthrobacter
  • Glycosyl Hydrolase Family
  • Intracellular Fraction
  • Arthrobacter Strain