title: Characterization of the N-acetylneuraminic acid synthase (NeuB) from the psychrophilic fish pathogen Moritella viscosa.
abstract: Moritella viscosa is a Gram-negative psychrophilic bacterium that causes winter ulcer disease in Atlantic salmon and cod. Its genome reveals that it possesses the ability to synthesize sialic acids. Indeed, sialic acid can be isolated from the bacterium and when analyzed using HPLC-MS/MS, the presence of N-acetylneuraminic acid was confirmed. Thus, the N-acetylneuraminic acid synthase NeuB from M. viscosa (MvNeuB) was recombinantly produced and characterized. The optimum pH and temperature for MvNeuB activity are 7.5 and 30 °C, respectively. The KM for N-acetylmannosamine and phosphoenolpyruvate is 18±5 and 0.8±0.2 mM, respectively. The kcat value (∼225 min(-1)) for both N-acetylmannosamine and phosphoenolpyruvate is the highest turnover number found for an enzyme in this class until the date. A calorimetric study of MvNeuB shows that the enzyme has a two-step transition peak probably reflecting the two domains these proteins consist of. MvNeuB is less stable at higher temperature and has a high catalytic activity at lower temperature compared to mesophilic counterparts. Enzymes from psychrophilic organisms are generally cold adapted meaning they can maintain adequate function near the freezing point of water. Cold adapted enzymes are catalytically more efficient at lower temperature and are more thermo-labile compared to their mesophilic counterparts. MvNeuB is a typical cold adapted enzyme and could be further explored for production of sialic acids and derivates at low temperatures.
authors.affiliation: The Norwegian Structural Biology Center (NorStruct), Department of Chemistry, University of Tromsø-The Arctic University of Norway, N-9037 Tromsø, Norway.
authors.forename: Tor Olav
authors.initials: TO
authors.lastname: Berg
authors.affiliation: The Norwegian Structural Biology Center (NorStruct), Department of Chemistry, University of Tromsø-The Arctic University of Norway, N-9037 Tromsø, Norway.
authors.forename: Man Kumari
authors.initials: MK
authors.lastname: Gurung
authors.affiliation: The Norwegian Structural Biology Center (NorStruct), Department of Chemistry, University of Tromsø-The Arctic University of Norway, N-9037 Tromsø, Norway.
authors.forename: Bjørn
authors.initials: B
authors.lastname: Altermark
authors.affiliation: The Norwegian Structural Biology Center (NorStruct), Department of Chemistry, University of Tromsø-The Arctic University of Norway, N-9037 Tromsø, Norway.
authors.forename: Arne O
authors.initials: AO
authors.lastname: Smalås
authors.affiliation: The Norwegian Structural Biology Center (NorStruct), Department of Chemistry, University of Tromsø-The Arctic University of Norway, N-9037 Tromsø, Norway. Electronic address: inger.l.rader@uit.no.
authors.forename: Inger Lin U
authors.initials: IL
authors.lastname: Ræder
mesh_terms: D000595:Amino Acid Sequence; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008670:Metals / Q000494:pharmacology; D008969:Molecular Sequence Data; D034221:Moritella / Q000201:enzymology*; D009154:Mutation; D007652:Oxo-Acid-Lyases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D055503:Protein Multimerization; D020836:Protein Structure, Quaternary; D017421:Sequence Analysis; D013696:Temperature
chemical_list: D008670:Metals; C018730:N-acetylneuraminate synthase; D007652:Oxo-Acid-Lyases
keywords: Cold active; Moritella viscosa; N-Acetylneuraminic acid synthase; NeuB; Sialic acid synthase