Gene cloning, recombinant expression, purification and characterization of l-methionine decarboxylase from Streptomyces sp. 590

Masaya Hayashi, Akane Okada, Kumiko Yamamoto, Tomomi Okugochi, Chika Kusaka, Daizou Kudou, Michiko Nemoto, Junko Inagaki, Yuu Hirose, Toshihide Okajima, Takashi Tamura, Kenji Soda, Kenji Inagaki

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

L-Methionine decarboxylase (MetDC) from Streptomyces sp. 590 depends on pyridoxal 5′-phosphate and catalyzes the non-oxidative decarboxylation of L-methionine to produce 3-methylthiopropylamine and carbon dioxide. MetDC gene (mdc) was determined to consist of 1,674 bp encoding 557 amino acids, and the amino acid sequence is similar to that of L-histidine decarboxylases and L-valine decarboxylases from Streptomyces sp. strains. The mdc gene was cloned and recombinant MetDC was heterologously expressed by Escherichia coli. The purification of recombinant MetDC was carried out by DEAE-Toyopearl and Ni-NTA agarose column chromatography. The recombinant enzyme was homodimeric with a molecular mass of 61,000 Da and showed optimal activity between 45 to 55 °C and at pH 6.6, and the stability below 30 °C and between pH 4.6 to 7.0. L-Methionine and L-norleucine were good substrates for MetDC. The Michaelis constants for L-methionine and L-norleucine were 30 and 73mM, respectively. The recombinant MetDC (0.50 U/ml) severely inhibited growth of human tumour cells A431 (epidermoid ovarian carcinoma cell line) and MDA-MB-231 (breast cancer cell line), however showed relatively low cytotoxicity for human normal cell NHDF-Neo (dermal fibroblast cell line from neonatal foreskin). This study revealed the properties of the gene and the protein sequence of MetDC for the first time.

Original languageEnglish
Pages (from-to)389-398
Number of pages10
JournalJournal of biochemistry
Volume161
Issue number4
DOIs
Publication statusPublished - Apr 1 2017

Keywords

  • L-methionine decarboxylase
  • antitumour enzyme
  • gene cloning
  • pyridoxal 50-phosphate
  • recombinant expression

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

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