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Deduced amino-acid sequence of a calcium-free α-amylase from a strain of Bacillus

Implications from molecular modeling of high oxidation stability and chelator resistance of the enzyme

  1. Hiroshi Hagihara,
  2. Yasuhiro Hayashi,
  3. Keiji Endo,
  4. Kazuaki Igarashi,
  5. Tadahiro Ozawa,
  6. Shuji Kawai,
  7. Katsuya Ozaki,
  8. Susumu Ito

Article first published online: 20 DEC 2001

DOI: 10.1046/j.1432-1327.2001.02308.x

Issue

European Journal of Biochemistry

European Journal of Biochemistry

Volume 268, Issue 14, pages 3974–3982, July 2001

Additional Information

How to Cite

Hagihara, H., Hayashi, Y., Endo, K., Igarashi, K., Ozawa, T., Kawai, S., Ozaki, K. and Ito, S. (2001), Deduced amino-acid sequence of a calcium-free α-amylase from a strain of Bacillus. European Journal of Biochemistry, 268: 3974–3982. doi: 10.1046/j.1432-1327.2001.02308.x

Author Information

  1. Tochigi Research Laboratories of Kao Corporation, Ichikai, Haga, Japan

* Y. Hayashi, Tochigi Research Laboratories of Kao Corporation, 2606 Akabane, Ichikai, Haga, Tochigi 321-3497, Japan. Fax: + 81 285687547, Tel.: + 81 285687516, E-mail: 304460@kastanet.kao.co.jp

Publication History

  1. Issue published online: 20 DEC 2001
  2. Article first published online: 20 DEC 2001
  3. (Received 5 March 2001, accepted 22 May 2001)

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Keywords:

  • α-amylase;
  • Bacillus;
  • amino-acid sequence;
  • molecular modeling;
  • oxidative stability

Alkaline α-amylase (AmyK38) from the alkaliphilic Bacillus sp. strain KSM-K38 is a unique enzyme in that it is highly chelator-resistant and oxidatively stable [Hagihara, H., Igarashi, K., Hayashi, Y., Endo, K., Ikawa-Kitayama, K., Ozaki, K., Kawai, S. & Ito, S. (2001) Appl. Environ. Microbiol.67, 1744–1750]. This enzyme was found to contain no Ca and require Na (or monovalent cations) for manifestation of activity. The nucleotide sequence of the gene for the novel enzyme was determined, and it harbored an ORF of 1503 bp encoding the enzyme of 501 amino acids, including a 21-amino-acid signal peptide. The deduced amino-acid sequence of the mature enzyme (55 097 Da) showed moderate homology to those of α-amylases from Bacillus licheniformis, Bacillus stearothermophilus and Bacillus amyloliquefaciens, with ≈ 63% identity. A methionine residue, which is conserved and susceptible to chemical oxidation, was replaced with leucine in AmyK38. Moreover, many conserved residues that are crucial ligands for Ca were replaced with other amino acids, thereby leading to loss of the Ca coordination geometries. By building a molecular model, we showed the calcium-independent, oxidatively stable active-site topology and structural integrity of AmyK38.

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