A novel mechanism of protein thermostability: a unique N-terminal domain confers heat resistance to Fe/Mn-SODs

Abstract

Superoxide dismutases (SODs), especially thermostable SODs, are widely applied in medical treatments, cosmetics, food, agriculture, and other industries given their excellent antioxidant properties. A novel thermostable cambialistic SOD from Geobacillus thermodenitrificans NG80-2 exhibits maximum activity at 70°C and high thermostability over a broad range of temperatures (20–80°C). Unlike other reported SODs, this enzyme contains an extra repeat-containing N-terminal domain (NTD) of 244 residues adjacent to the conserved functional SODA domain. Deletion of the NTD dramatically decreased its optimum active temperature (OAT) to 30°C and also impaired its thermostability. Conversely, appending the NTD to a mesophilic counterpart from Bacillus subtilis led to a moderately thermophilic enzyme (OAT changed from 30 to 55°C) with improved heat resistance. Temperature-dependant circular dichroism analysis revealed the enhanced conformational stability of SODs fused with this NTD. Furthermore, the NTD also contributes to the stress resistance of host proteins without altering their metal ion specificity or oligomerisation form except for a slight effect on their pH profile. We therefore demonstrate that the NTD confers outstanding thermostability to the host protein. To our knowledge, this is the first discovery of a peptide capable of remarkably improving protein thermostability and provides a novel strategy for bioengineering thermostable SODs.

Unique Characteristics of Recombinant Hybrid Manganese Superoxide Dismutase from Staphylococcus equorum and S. saprophyticus

“…Interestingly, such stabilization can be achieved by introducing a charged amino acid residue at only the N-or C-termini [44]. The N-terminal domain of Fe/Mn-SOD from Geobacillus thermodenitrificans NG80-2 has been reported to be responsible for the enzyme thermostability, as well as appending thermostability to mesophilic Fe/Mn-SOD from Bacillus subtilis upon fusion [11]. The N-terminal part of rMnSODSeq is constructed based on the amino acid sequence of that of S. saprophyticus, which not resembles the N-terminal domain of G. thermodenitrificans NG80-2.…”

“…Interestingly, few SODs can also be cambialistic, which means that the type of metal installed is exchangeable [20]. Further, SODs demonstrate stability against various environmental stress, particularly those originated from thermophilic or hyperthermophilic microorganisms [8][9][10][11][12][13][14]21]. Thermostable SOD are of interest for application such as sun cream and for its stability during production, formulation, shipping, and storage.…”

“…Many SODs are reported to display low activity at low pH [7][8][9][10][11][12][13][14] and to cope with that, strategies have been developed to improve the enzyme stability towards extreme pH condition. Immobilization the enzyme to nanoparticles appears as an attractive alternative because it prolongs and extends the use of the enzyme, as demonstrated in stabilization of SOD from Thermus thermophillus wl [15].…”

Inorganic salts and compatible solutes help mesophilic bacteria inhabit the high temperature waters of a Trans-Himalayan sulfur-borax spring

“…Furthermore, environment-aided survival of mesophiles in high temperature habitats seems feasible in the light of the fact that researchers have found no special molecular/structural 85 biological feature in the biomacromolecules of thermophiles/hyperthermophiles that can be exclusively attributed to their high temperature adaptation. In other words, all the biophysical features which have thus far been attributed to the high-temperature adaptation of thermophiles/hyperthermophiles (via in vitro studies)for instance specialized topologies and functions of proteins, nucleic acids and membrane lipids (Kumar et al, 2000;Dekker et al, 2003, Zeldovich et al, 2007Koga, 2012;Wang et al, 2014), protein-stabilizing adaptive 90 mutations (Fields, 2001), and presence of compatible solutes (Welsh, 2000;Fields, 2001) -are present in some mesophile or the other (Fields, 2001;Vieille and Zeikus, 2001;Hallsworth et al, 2003;Rudolph et al, 2010;Dibrova et al, 2014;Ezemaduka et al, 2014;de Lima Alves et al, 2015;Hamerly et al, 2015;Jing et al, 2017;Pucci and Rooman, 2017).…”

The first crystal structure of manganese superoxide dismutase from the genusStaphylococcus

“…However, these homologous enzymes have significantly different characteristics. For example, S. equorum MnSOD is still active at 353 K (Retnoningrum et al, 2016), at which temperature B. subtilis MnSOD is completely inactivated (Wang et al, 2014). S. equorum MnSOD is also more resistant to inactivation by urea or guanidinium chloride (chaotropic agents), by sodium dodecyl sulfate (a detergent) or by -mercaptoethanol (a reducing agent) than B. subtilis MnSOD (Retnoningrum et al, 2016).…”