Highly Stable, Cold-Active Aldehyde Dehydrogenase from the Marine Antarctic Flavobacterium sp. PL002

Georgiana Necula-Petrareanu,Paris Lavin,Victoria Ioana Paun,Giulia Roxana Gheorghita,Cristina Purcarea,Alina Vasilescu

doi:10.3390/fermentation8010007

Abstract

Stable aldehyde dehydrogenases (ALDH) from extremophilic microorganisms constitute efficient catalysts in biotechnologies. In search of active ALDHs at low temperatures and of these enzymes from cold-adapted microorganisms, we cloned and characterized a novel recombinant ALDH from the psychrotrophic Flavobacterium PL002 isolated from Antarctic seawater. The recombinant enzyme (F-ALDH) from this cold-adapted strain was obtained by cloning and expressing of the PL002 aldH gene (1506 bp) in Escherichia coli BL21(DE3). Phylogeny and structural analyses showed a high amino acid sequence identity (89%) with Flavobacterium frigidimaris ALDH and conservation of all active site residues. The purified F-ALDH by affinity chromatography was homotetrameric, preserving 80% activity at 4 °C for 18 days. F-ALDH used both NAD+ and NADP+ and a broad range of aliphatic and aromatic substrates, showing cofactor-dependent compensatory KM and kcat values and the highest catalytic efficiency (0.50 µM−1 s−1) for isovaleraldehyde. The enzyme was active in the 4–60 °C-temperature interval, with an optimal pH of 9.5, and a preference for NAD+-dependent reactions. Arrhenius plots of both NAD(P)+-dependent reactions indicated conformational changes occurring at 30 °C, with four(five)-fold lower activation energy at high temperatures. The high thermal stability and substrate-specific catalytic efficiency of this novel cold-active ALDH favoring aliphatic catalysis provided a promising catalyst for biotechnological and biosensing applications.

Highlights

Extremophilic bacteria are characterized by tolerance or the capability of growing under extreme conditions, such as hypersaline habitats, high pressures, and extreme temperatures
We report on the cloning, heterologous expression, and biochemical characterization of a cold-active aldehyde dehydrogenases (ALDH) from the Antarctic alga-associated Flavobacterium sp
The phylogenetic analysis (Figure 1) of the F-ALDH amino acid sequence in relation with the homologous aldehyde dehydrogenases from Flavobacterium spp. and other genera showed the Antarctic enzyme grouped with ALDSs from the Clade I of the genus Flavobacterium

Summary

Introduction

Extremophilic bacteria are characterized by tolerance or the capability of growing under extreme conditions, such as hypersaline habitats, high pressures, and extreme temperatures. Many extremophiles inhabiting environments characterized by multiple extreme conditions are known as polyextremophiles [1,2]. The capability to overcome such stressing conditions present in extreme environments, is due to special characteristics of their extremophilic enzymes (extremozymes) [3,4,5]. Research has exploited extreme environments looking for producers of a wide array of biotechnologically useful extremozymes that are one of the great interests for industrial processes, mainly in biocatalysis due to their particular stability and high catalytic activity over a wide range of various parameters (temperature, salinity, pH, etc.).

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Conclusion