An Azide-Insensitive Superoxide Dismutase from a Hyperthermophilic Archaeon, Sulfolobus solfataricus

Abstract

The superoxide dismutase (SOD) gene of Sulfolobus solfataricus, a hyperthermophilic archaeon, was cloned and expressed in Escherichia coli, and its gene product was characterized. When the protein was expressed in E. coli, it formed a homodimer that contained both Mn and Fe. Metal reconstitution experiments of the SOD with Fe or Mn showed that only the Fe-reconstituted SOD was active. Substitution of Tyr88 to Phe did not affect the metal specificity of the enzyme. The Fe-reconstituted SOD was extremely resistant to thermal denaturation; e.g. 96% of the initial activity was retained after heating at 95 degreesC for 2 h. Fe-reconstituted SOD was not inhibited by azide, but fluoride inhibition was observed. This suggests that some steric hindrance in the substrate funnel of the enzyme prevents the access of N3- but allows O2- and F- access to the active site.

Biochemical Properties and Regulated Gene Expression of the Superoxide Dismutase from the Facultatively Aerobic Hyperthermophile Pyrobaculum calidifontis

“…Both enzymes displayed relatively strong resistance against azide (Mnand Fe-SOD activity decreased 50% with 380 and 340 mM azide, respectively). This resistance against azide has also been observed in other archaeal SODs (44,45).…”

“…Mn-and Fe-SODs are closely related in amino acid sequence and three-dimensional structure. Among (hyper)thermophilic archaea, Fe-SODs have been reported from Sulfolobus solfataricus (11,44), Sulfolobus acidocaldarius (18), Acidianus ambivalens (17), and Thermoplasma acidophilum (37). Although Mn-and Fe-SODs generally exhibit a strict metal specificity for SOD activity (34), several cambialistic SODs, which are active with either Mn or Fe, have been reported.…”

Protection of Methanosarcina barkeri against oxidative stress: identification and characterization of an iron superoxide dismutase

“…Superoxide dismutase from M. barkeri was inhibited by azide and H 2 O 2 but to a lessr extent than the iron superoxide dismutase from E. coli (Sigma), which we used as control (Table 2). A relative insensitivity to azide and H 2 O 2 has also been reported for the iron superoxide dismutases from Methanobacterium thermoautotrophicum and Sulfolobus solfataricus (Takao et al 1991;Yamano and Maruyama 1999). Superoxide dismutase from M. barkeri was not inhibited by 10 mM cyanide, which is a property shared among all iron and manganese superoxide dismutases.…”

A change of the metal-specific activity of a cambialistic superoxide dismutase from Porphyromonas gingivalis by a double mutation of Gln-70 to Gly and Ala-142 to Gln

“…Although a few exceptions have been reported [17,18], these three differences could be the primary candidates to account for the metal specificity of the Fe-and Mn-SOD activity [19], improving on the suggestions by Parker and Blake [20] and Yamakura et al [21]. Recently, Yamano and Maruyama [22] reported that the substitution of Tyr-77 by Phe in the metal-specific Fe-SOD from Sulfolobus solfataricus did not change the metal-specific activity of the enzyme. Therefore, the third difference may not contribute to the metal-specific activity of Fe-and Mn-SODs.…”