Azoreductase from alkaliphilic Bacillus sp. AO1 catalyzes indigo reduction

Hirokazu Suzuki, Tomoaki Abe, Katsumi Doi, Toshihisa Ohshima

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

Indigo is an insoluble blue dye historically used for dyeing textiles. A traditional approach for indigo dyeing involves microbial reduction of polygonum indigo to solubilize it under alkaline conditions; however, the mechanism by which microorganisms reduce indigo remains poorly understood. Here, we aimed to identify an enzyme that catalyzes indigo reduction; for this purpose, from alkaline liquor that performed microbial reduction of polygonum indigo, we isolated indigo carmine-reducing microorganisms. All isolates were facultative anaerobic and alkali-tolerant Bacillus spp. An isolate termed AO1 was found to be an alkaliphile that preferentially grows at pH 9.0–11.0 and at 30–35 °C. We focused on flavin-dependent azoreductase as a possible enzyme for indigo carmine reduction and identified its gene (azoA) in Bacillus sp. AO1 using homology-based strategies. azoA was monocistronic but clustered with ABC transporter genes. Primary sequence identities were < 50% between the azoA product (AzoA) and previously characterized flavin-dependent azoreductases. AzoA was heterologously produced as a flavoprotein tolerant to alkaline and organic solvents. The enzyme efficiently reduced indigo carmine in an NADH-dependent manner and showed strict specificity for electron acceptors. Notably, AzoA oxidized NADH in the presence, but not the absence, of indigo. The reaction rate was enhanced by adding organic solvents to solubilize indigo. Absorption spectrum analysis showed that indigo absorption decreased during the reaction. These observations suggest that AzoA can reduce indigo in vitro and potentially in Bacillus sp. AO1. This is the first study that identified an indigo reductase, providing a new insight into a traditional approach for indigo dyeing.

Original languageEnglish
Pages (from-to)9171-9181
Number of pages11
JournalApplied Microbiology and Biotechnology
Volume102
Issue number21
DOIs
Publication statusPublished - Nov 1 2018

Fingerprint

Indigo Carmine
Bacillus
Polygonum
azoreductase
NAD
Enzymes
Flavoproteins
ATP-Binding Cassette Transporters
Textiles
Alkalies

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Applied Microbiology and Biotechnology

Cite this

Azoreductase from alkaliphilic Bacillus sp. AO1 catalyzes indigo reduction. / Suzuki, Hirokazu; Abe, Tomoaki; Doi, Katsumi; Ohshima, Toshihisa.

In: Applied Microbiology and Biotechnology, Vol. 102, No. 21, 01.11.2018, p. 9171-9181.

Research output: Contribution to journalArticle

Suzuki, Hirokazu ; Abe, Tomoaki ; Doi, Katsumi ; Ohshima, Toshihisa. / Azoreductase from alkaliphilic Bacillus sp. AO1 catalyzes indigo reduction. In: Applied Microbiology and Biotechnology. 2018 ; Vol. 102, No. 21. pp. 9171-9181.
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