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

Hiraoka, B. Yukihiro; Yamakura, Fumiyuki; Sugio, Shigetoshi; Nakayama, Koji

doi:10.1042/bj3450345

Cited by 34 publications

(7 citation statements)

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“…Despite recent reports of changes in metal specificity in engineered SOD enzymes [31,32], the results have been unspectacular, resulting in small increases in enzyme activity in SOD holoenzymes reconstituted with the ‘wrong’ metal. Unfortunately, the H145E mutation of Mycobacterium tuberculosis FeSOD which produces an active MnSOD [49] bears no relevance to the central question of specificity as no other SODs with this mutation exist naturally.…”

Section: Discussionmentioning

confidence: 99%

“…Double mutations have been introduced into P. gingivalis and E. coli SODs with the intention of altering their metal specificities. In the former cambialistic enzyme, mutations Q70G/A142Q reduced the iron‐supported activity of the enzyme and altered the ratio of Mn : Fe from 1.4 to 3.5 [31]. In the latter MnSOD, the equivalent mutation G77Q/Q146A was demonstrated to reduce specific activity to 71% and introduce an iron‐supported SOD activity which did not exist in the wild‐type enzyme, though this was only 6% of manganese‐supported activity in the wild type [32].…”

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confidence: 99%

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Thermostability of manganese‐ and iron‐superoxide dismutases from Escherichia coli is determined by the characteristic position of a glutamine residue

Hunter¹,

Bannister²,

Hunter³

2002

European Journal of Biochemistry

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The structurally homologous mononuclear iron and manganese superoxide dismutases (FeSOD and MnSOD, respectively) contain a highly conserved glutamine residue in the active site which projects toward the active-site metal centre and participates in an extensive hydrogen bonding network. showed reduced selectivity toward their metal. Differences exhibited in the thermostability of SOD activity was most obvious in the mutants that contained two glutamine residues (FeSOD[A141Q] and MnSOD[G77Q]), where the MnSOD mutant was thermostable and the FeSOD mutant was thermolabile. Significantly, the MnSOD double mutant exhibited a thermal-inactivation profile similar to that of wild-type FeSOD while that of the FeSOD double mutant was similar to wild-type MnSOD. We conclude therefore that the position of this glutamine residue contributes to metal selectivity and is responsible for some of the different physicochemical properties of these SODs, and in particular their characteristic thermostability.Keywords: superoxide dismutase; site-directed mutagenesis; metal specificity; thermostability.Iron superoxide dismutases (FeSOD, E.C.1.15.1.1) and manganese superoxide dismutases (MnSOD) constitute a class of structurally equivalent metalloenzymes prevalent in prokaryotes and in eukaryotic mitochondria, respectively. They exhibit a very high degree of homology in both sequence and structure (Fig. 1). The SODs are active only in dimeric association and all share structural homology in this respect [1]. The metal cofactors are required to catalyse the disproportionation of the superoxide radical into hydrogen peroxide and molecular oxygen [2] in a cyclic, two-stage oxidation-reduction mechanism:where M represents either iron or manganese. Selectivity of the proteins for their metal cofactor has been demonstrated in vivo [3] and although apoenzymes of each type of SOD may be reconstituted by the addition of metals, the resulting enzyme is active only with the authentic metal at its active centre [4][5][6][7]. A small number of cambialistic SODs have been shown to be active with either iron or manganese, though only those of Propionibacterium shermanii [8], Bacteroides gingivalis [9] and Bacteroides fragilis [10] demonstrate similar specific activities with either metal.In all structures solved for the mononuclear SODs, the metal ion is held in place by an absolutely conserved quartet of residues comprising three histidines and one aspartic acid which act as ligands to the metal (H26, H81, D167 and H171 for Escherichia coli MnSOD, Fig. 1B. This numbering will be used throughout except where indicated) [11][12][13][14][15][16][17][18][19][20][21]. This conservation is also reflected in all sequences elucidated for this large group of ubiquitous enzymes. A fifth metal ligand, either water or hydroxide, present in all structures produces a trigonal-pyramidal geometry at the active site. A distorted-octahedral geometry is assumed during catalytic turnover or inhibition,

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Thermostability of manganese‐ and iron‐superoxide dismutases from Escherichia coli is determined by the characteristic position of a glutamine residue

Hunter¹,

Bannister²,

Hunter³

2002

European Journal of Biochemistry

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“…On the other hand, no structural interpretation for the bases of these two chemical mechanisms has been proposed yet. Recently, we [41] have made a mutant of P. gingivalis SOD, which has complementary substitution of Gln70 with Gln142, that is from Fe-SOD type Gln-centered hydrogen-bond network to the Mn-SOD type. This mutant shifed its metal-specific activity from almost equally efficient with both Fe and Mn to dominantly Mn efficient.…”

Section: Metals-specific Activitymentioning

confidence: 99%

Crystal structure of cambialistic superoxide dismutase from Porphyromonas gingivalis

Sugio

Hiraoka

Yamakura

2000

European Journal of Biochemistry

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The crystal structure of cambialistic superoxide dismutase (SOD) from Porphyromonas gingivalis, which exhibits full activity with either Fe or Mn at the active site, has been determined at 1.8-A Ê resolution by molecular replacement and refined to a crystallographic R factor of 17.9% (R free 22.3%). The crystals belong to the space group P2 1 2 1 2 1 (a 75.5 A Ê , b 102.7 A Ê , c 99.6 A Ê ) with four identical subunits in the asymmetric unit. Each pair of subunits forms a compact dimer, but not a tetramer, with 222 point symmetry. Each subunit has 191 amino-acid residues most of which are visible in electron density maps, and consists of seven a helices and one three-stranded antiparallel b sheet. The metal ion, a 3 : 1 mixture of Fe and Mn, is coordinated with five ligands (His27, His74, His161, Asp157, and water) arranged at the vertices of a trigonal bipyramid. Although the overall structural features, including the metal coordination geometry, are similar to those found in other single-metal containing SODs, P. gingivalis SOD more closely resembles the dimeric Fe-SODs from Escherichia coli rather than another cambialistic SOD from Propionibacterium shermanii, which itself is rather similar to other tetrameric SODs.Keywords: crystal structure; cambialistic superoxide dismutase; iron superoxide dismutase; manganese superoxide dismutase.Superoxide dismutase (SOD) is an enzyme catalyzing disproportionation of the superoxide radical to hydrogen peroxide and dioxygen by the following scheme: Mn is the majority of the bound metal species [16]. Despite its cambialistic nature, the protein has a very similar polypeptide chain folding and virtually identical spatial arrangement of the active site residues to other metal-specific SODs. As a thorough comparison of Fe-and Mn-substituted SODs does not provide significant structural differences [16], the structural basis of cambialism of Pr. shermanii SOD is still unclear. SOD from P. gingivalis is also cambialistic; it intrinsically contains both iron and manganese at a different ratio depending on differences in air supply to the culture [15]. Yamakura et al. have shown that a recombinant P. gingivalis SOD can be reconstituted with either Fe or Mn exclusively, and both the Fe-and Mn-substituted enzymes showed almost the same specific activities as the native enzyme [17].Recent structural studies suggest that cambialistic SODs could be divided into two types according to the similarity of their primary structures and the difference in the hydrogenbonding network close to the active-site metals. The primary structure of Pr. shermanii SOD is closer to that of terameric Fe-SOD from Mycobacterium tuberculosis and Mn-SODs [18], while the primary structure of P. gingivalis SOD more closely resembles that of dimeric Fe-SODs from Escherichia coli and Pseudomonas ovalis [19]. Hydrogen-bond networks around the active-site metal are constituted differently between the two cambialistic SODs. In Pr. shermanii SOD, His142 (amino-acid numbering is based on the positions in P. gingivalis...

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“…Gram-negative bacteria that use a switch-like riboregulatory mechanism are depicted with dashed lines. In addition to the species mentioned in the text, the following bacterial species that code for SOD enzymes are shown in the figure: Neisseria gonorrhoeae and N. meningitidis (Archibald and Duong, 1986;Seib et al, 2004), Haemophilus influenzae (Kroll et al, 1998), Porphyromonas gingivalis (Hiraoka et al, 2000), Coxiella burnetii (Heinzen et al, 1992), Rickettsia typhi (McLeod et al, 2004), Burkholderia pseudomallei (Loprasert et al, 2000), Campylobacter jejuni (Atack and Kelly, 2009), Legionella pneumophila (Sadosky et al, 1994), Francisella tularensis (Bakshi et al, 2006), Helicobacter pylori (Ernst et al, 2005), Chlamydia pneumoniae (Yu et al, 2004), Borrelia burgdorferi (Aguirre et al, 2013), Brucella abortus (Sriranganathan et al, 1991), Streptococcus pneumoniae, S. mutans, and S. pyogenes (Gerlach et al, 1998;De Vendittis et al, 2010;Eijkelkamp et al, 2014), Mycobacterium tuberculosis and M. smegmatis (Yamakura et al, 1995;Bunting et al, 1998;Padilla-Benavides et al, 2013), Corynebacterium diphtheriae (Britton et al, 1978), and C. glutamicum (El Shafey et al, 2008), Listeria monocytogenes (Britton et al, 1978) Enterococcus faecalis (Verneuil et al, 2006;Wasselin et al, 2021), Nocardia asteroides (Beaman et al, 1983), and Deinococcus radiophilus (Yun and Lee, 2004).…”

Section: Mn-sod Requires That Bacteria Facilitate Its Correct Metalla...mentioning

confidence: 99%

Why is manganese so valuable to bacterial pathogens?

Čapek

Večerek

2023

Front. Cell. Infect. Microbiol.

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Apart from oxygenic photosynthesis, the extent of manganese utilization in bacteria varies from species to species and also appears to depend on external conditions. This observation is in striking contrast to iron, which is similar to manganese but essential for the vast majority of bacteria. To adequately explain the role of manganese in pathogens, we first present in this review that the accumulation of molecular oxygen in the Earth’s atmosphere was a key event that linked manganese utilization to iron utilization and put pressure on the use of manganese in general. We devote a large part of our contribution to explanation of how molecular oxygen interferes with iron so that it enhances oxidative stress in cells, and how bacteria have learned to control the concentration of free iron in the cytosol. The functioning of iron in the presence of molecular oxygen serves as a springboard for a fundamental understanding of why manganese is so valued by bacterial pathogens. The bulk of this review addresses how manganese can replace iron in enzymes. Redox-active enzymes must cope with the higher redox potential of manganese compared to iron. Therefore, specific manganese-dependent isoenzymes have evolved that either lower the redox potential of the bound metal or use a stronger oxidant. In contrast, redox-inactive enzymes can exchange the metal directly within the individual active site, so no isoenzymes are required. It appears that in the physiological context, only redox-inactive mononuclear or dinuclear enzymes are capable of replacing iron with manganese within the same active site. In both cases, cytosolic conditions play an important role in the selection of the metal used. In conclusion, we summarize both well-characterized and less-studied mechanisms of the tug-of-war for manganese between host and pathogen.

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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

Cited by 34 publications

References 30 publications

Thermostability of manganese‐ and iron‐superoxide dismutases from Escherichia coli is determined by the characteristic position of a glutamine residue

Thermostability of manganese‐ and iron‐superoxide dismutases from Escherichia coli is determined by the characteristic position of a glutamine residue

Crystal structure of cambialistic superoxide dismutase from Porphyromonas gingivalis

Why is manganese so valuable to bacterial pathogens?

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