Abstract

Psychrophilic bacteria, extremophiles that thrive at low temperatures, contain enzymes adapted to function in cold environments. NAD+ -dependent aldehyde dehydrogenase (F-ALDH) from Flavobacterium PL02, an Antarctic isolated bacterial strain, and the aspartate transcarbamoylase (ATC) from Glaciibacter superstes originating from an Alaska ice brine were investigated as catalyst in biosensing and cold adaptation biomarker, respectively. The protein sequence of these cold-active enzymes is homologous to corresponding proteins from other psychrophlic and psychrotrophic bacteria. Structural analysis revealed in both cases a specific aminoacid composition favouring the protein flexibility under low temperatures such as the reduced number of cysteine residues. 3D modelling indicated insertions of loop-motifs both at subunit interfaces and at the C-terminus, providing enzyme flexibility to performed catalysis at near freezing temperatures. Hydrophobic core analysis revealed the distribution of reduced hydrophobic cores as compared to mesophilic and thermophilic counterparts related to protein stability. Cloning and functional characterization of F-ALDH expressed in E. coli provided a stable and active catalyst for aldehyde oxidation at temperatures as low as 4ºC, capable of using a series of aliphatic and aromatic substrates, providing a valuable biocomponent for cold-active biosensors for wine, cosmetics and food industries. The quantitative response of G. superstes ATC exposed to heat-shock cycles as a model experiment for microbial response to environmental changes indicated this cold-adapted enzyme as a valuable marker for bacterial resilience.