The first structure of a cold-adapted superoxide dismutase (SOD): biochemical and structural characterization of iron SOD fromAliivibrio salmonicida

Abstract

Superoxide dismutases (SODs) are metalloenzymes that catalyse the dismutation of the superoxide radical anion into O(2) and H(2)O(2) in a two-step reaction. The crystal structure of the iron superoxide dismutase from the cold-adapted and fish-pathogenic bacterium Aliivibrio salmonicida (asFeSOD) has been determined and refined to 1.7 A resolution. The protein has been characterized and compared with the closely related homologous iron superoxide dismutase from the mesophilic Escherichia coli (ecFeSOD) in an attempt to rationalize its environmental adaptation. ecFeSOD shares 75% identity with asFeSOD. Compared with the mesophilic FeSOD, the psychrophilic FeSOD has distinct temperature differences in residual activity and thermostability that do not seem to be related to structural differences such as intramolecular or intermolecular ion bonds, hydrogen bonds or cavity sizes. However, an increased net negative charge on the surface of asFeSOD may explain its lower thermostability compared with ecFeSOD. Activity measurements and differential scanning calorimetry measurements revealed that the psychrophilic asFeSOD had a thermostability that was significantly higher than the optimal growth temperature of the host organism.

Structure and flexibility in cold-adapted iron superoxide dismutases: The case of the enzyme isolated from Pseudoalteromonas haloplanktis

“…The overall structure of PhSOD is very close to that of EcSOD (Lah et al, 1995), PoSOD (Ringe et al, 1983), and AsSOD (Pedersen et al, 2009), thus showing that the differences in the amino acid sequence do not produce any relevant alteration in the tertiary structure. Indeed, after superimposition of PhSOD on the other three structures, the RMSD of all the Ca atoms is in the range 0.45-0.54 Å.…”

“…2A). The metal coordination is strictly conserved among all Fe-and Mn-SODs, whose structures have been determined so far (Boucher et al, 2006;Boucher et al, 2005;Lah et al, 1995;Munoz et al, 2005;Pedersen et al, 2009;Ringe et al, 1983;Ursby et al, 1999). The geometry of the coordination is a slightly distorted trigonal bipyramid, in which His26 Ne2 and the oxygen of the water sit at the axial top of each pyramid, while Ne2 atoms of His73 and His161, and Od1 of Asp157 are the equatorial ligands.…”

“…Its polypeptide chain contains 192 residues, corresponding to a molecular mass of 21,251 Da. Paired sequence alignments indicated that PhSOD shares 77% amino acid identity with Fe-SOD from the cold-adapted bacterium Aliivibrio salmonicida (As-SOD), whose crystal structure was recently determined (Pedersen et al, 2009), and 69% amino acid identity with the mesophilic counterparts from Escherichia coli (EcSOD) (Lah et al, 1995) and Pseudomonas ovalis (PoSOD) (Ringe et al, 1983). Furthermore, 51% and 41% amino acid identity was found with the thermophilic SOD from Cyanobacterium thermosynecoccus elongatus (Kerfeld et al, 2003) and Sulfolobus solfataricus (SsSOD) (Dello Russo et al, 1997;Ursby et al, 1999), respectively.…”

PCR Amplification of a Superoxide Dismutase Gene (pasod) from Psychrophilic Bacteria (KOPRI22215) without Genome Information

“…7 Recently, the structure of a cold-adapted SOD was demonstrated with A. salmonicida, whose FeSOD exhibits lower thermostability compared with that of Escherichi coli (E. coli) probably due to its surface having an increased net negative charge. 8 It is of interest to us to search for cold-adapted SODs from Psychromonas arctica (KOPRI22215) 9 and compare them with those of E. coli. It is accordingly inferred that P. arctica might have developed an efficient protection system against oxidative stress resulting from reduced metabolic turnovers in respiration pathways.…”

Expression and Purification of Recombinant Superoxide Dismutase (PaSOD) from Psychromonas arctica in Escherichia coli

“…[17][18][19] Recently, the first crystal structure of FeSOD from the cold-adapted and fish-pathogenic bacterium Aliivibrio salmonicida (AsFeSOD) was determined. 20 Our group has previously reported the existence of SOD in P. arctica Abbreviations: SOD; superoxide dismutase, PCR; polymerase chain reaction, IPTG; isopropyl-β-D-1-thiogalactopyranoside, NBT; nitro blue tetrazolium, WST-1; 2-(4-iodophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfophenyl)-2H-tetrazolium (PaSOD), whose genomic sequence has not yet been elucidated. 18 We were able to identify the coding gene for sod from P. arctica using a combination of gradient PCR with homologous primers and LA PCR in vitro cloning technology.…”