Unique structural features of the monomeric Cu,Zn superoxide dismutase from Escherichia coli, revealed by X-ray crystallography

Pesce, Alessandra; Capasso, Clemente; Battistoni, Andrea; Folcarelli, Silvia; Rotilio, Giuseppe; Desideri, Alessandro; Bolognesi, Martino

doi:10.1006/jmbi.1997.1400

Cited by 81 publications

(69 citation statements)

References 49 publications

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“…In particular, the hydrophobic residues Phe-50 and Gly-51, essential for stabilizing a dimeric form (26), are present in prokaryotic SODs in only a few cases. In E. coli (9) and in B. subtilis (this article), which are monomeric in solution, there is a Lys at position 51. This charged group could be determinant for preventing occurrence of the quaternary structure.…”

Section: Resultsmentioning

confidence: 98%

“…SOD has been considered for a long time to be peculiar of eukaryotic organisms, in which its evolutionary rate has been studied (5). From the first description of a SOD activity in Escherichia coli (6), other putative homologous of the eukaryotic protein have been then found in the periplasmic space of other bacterial species (7) The x-ray structures have been solved for the proteins from Photobacterium leiognathi (8), E. coli (9), Salmonella typhimurium (10), and Actinobacillus pleuropneumoniae (11), all of them maintaining the typical metal sites of Cu,Zn eukaryotic SODs and normal SOD activity. With the aim of providing a more detailed picture of prokaryotic SODs, we browsed the available complete genomes of Archea and Bacteria to search for sequences with some degree of homology with human SOD (HSOD), which is found in 48 of the 138 prokaryotes with complete genome.…”

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A prokaryotic superoxide dismutase paralog lacking two Cu ligands: From largely unstructured in solution to ordered in the crystal

Banci

Bertini

Calderone

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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Little is known about prokaryotic homologs of Cu,Zn superoxide dismutase (SOD), an enzyme highly conserved among eukaryotic species. In 138 Archaea and Bacteria genomes, 57 of these putative homologs were found, 11 of which lack at least one of the metal ligands. Both the solution and the crystal structures of the SOD-like protein from Bacillus subtilis, lacking two Cu ligands and found to be enzymatically inactive, were determined. In solution, the protein is monomeric. The available nuclear Overhauser effects, together with chemical-shift index values, allowed us to define and to recognize the typical Cu,Zn SOD Greek ␤-barrel but with largely unstructured loops (which, therefore, sample a wide range of conformations). On the contrary, in the crystal structure (obtained in the presence of slight excess of Zn), the protein is well structured and organized in covalent dimers held by a symmetric bridge consisting of a Zn ion bound to an Asp-His dyad in a tetrahedral geometry. Couples of dimers held by hydrophobic interactions and H bonds are further organized in long chains. The order͞disorder transition is discussed in terms of metal binding and physical state.conformational mobility ͉ NMR structure determination ͉ Zn-mediated protein dimers ͉ x-ray crystallography ͉ superoxide dismutase T he availability of the complete sequence of an increasing number of genomes allows comparative analyses aimed at the identification of genes and of corresponding physiological functions conserved along evolution. Cu,Zn superoxide dismutase (SOD) is a ubiquitous family of enzymes that efficiently catalyze the dismutation of superoxide anions into oxygen and hydrogen peroxide (1, 2). The protein takes a classical Greek key ␤-barrel fold and adopts a dimeric quaternary structure (3). It binds one Zn and one Cu ion per subunit, the latter ion being the site of the enzymatic reaction, whose rate limiting step is the diffusion of superoxide through the channel that leads to the Cu site (4). SOD has been considered for a long time to be peculiar of eukaryotic organisms, in which its evolutionary rate has been studied (5). From the first description of a SOD activity in Escherichia coli (6), other putative homologous of the eukaryotic protein have been then found in the periplasmic space of other bacterial species (7) The x-ray structures have been solved for the proteins from Photobacterium leiognathi (8), E. coli (9), Salmonella typhimurium (10), and Actinobacillus pleuropneumoniae (11), all of them maintaining the typical metal sites of Cu,Zn eukaryotic SODs and normal SOD activity. With the aim of providing a more detailed picture of prokaryotic SODs, we browsed the available complete genomes of Archea and Bacteria to search for sequences with some degree of homology with human SOD (HSOD), which is found in 48 of the 138 prokaryotes with complete genome. In some of them, not all of the metal ligands are present. The SOD-like protein from Bacillus subtilis (BsSOD) lacks two binding-Cu residues (His-48 and His-63, human protein numbe...

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Section: Resultsmentioning

confidence: 98%

mentioning

confidence: 99%

A prokaryotic superoxide dismutase paralog lacking two Cu ligands: From largely unstructured in solution to ordered in the crystal

Banci

Bertini

Calderone

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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“…In particular, in the paper by Zhou et al (1997) it is shown that the effect of charge mutations on the association rate is predictable simply by calculating the enzyme-substrate interaction potential within the active site and that such effects are roughly proportional to the distance of the mutated site from the active site. In this context it is important to notice that bacterial Cu,Zn SODs, which have been shown to display high catalytic efficiency Foti et al, 1997;Folcarelli et al, 1998), lack the electrostatic loop containing the four charged residues neutralized in this work (Bourne et al, 1996;Pesce et al, 1997), whileArg141 is sequence invariant in all the Cu,Zn SODs known so far (Bordo et al, 1994). However, in the prokaryotic enzymes there is the insertion of a new loop, in the proximity of the active site, containing charged residues able to modulate the attraction of the superoxide anion, at least at low ionic strength (Folcarelli et al, 1998).…”

Section: Fig 2 Electrostatic Potential Distribution Around (A) Lqqtmentioning

confidence: 85%

Role of the electrostatic loop charged residues in Cu, Zn superoxide dismutase

et al. 1998

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We have expressed and characterized a mutant of Xenopus laevis Cu,Zn superoxide dismutase in which four highly conserved charged residues belonging to the electrostatic loop have been replaced by neutral side chains: Lysl20 + Leu, Asp130 + Gln, Glu131 + Gln, and Lys134 -+ Thr. At low ionic strength, the mutant enzyme is one of the fastest superoxide dismutases ever assayed (k = 6.7 X 10' M" s-' , at pH 7 and p = 0.02 M). Brownian dynamics simulations give rise to identical enzyme-substrate association rates for both wild-type and mutant enzymes, ruling out the possibility that enhancement of the activity is due to pure electrostatic factors. Comparative analysis of the experimental catalytic rate of the quadruple and single mutants reveals the nonadditivity of the mutation effects, indicating that the hyperefficiency of the mutant is due to a decrease of the energy barrier and/or to an alternative pathway for the diffusion of superoxide within the active site channel. At physiological ionic strength the catalytic rate of the mutant at neutral pH is similar to that of the wild-type enzyme as it is to the catalytic rate pH dependence. Moreover, mutation effects are additive. These results show that, at physiological salt conditions, electrostatic loop charged residues do not influence the diffusion pathway of the substrate and, if concomitantly neutralized, are not essential for high catalytic efficiency of the enzyme, pointing out the role of the metal cluster and of the invariant Arg 141 in determining the local electrostatic forces facilitating the diffusion of the substrate towards the active site.

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“…Compared with human SOD1, E. coli SodC possesses a seven-residue insertion in the loop region ("S-S subloop") containing Cys 74 , which is tethered with Cys 169 through the disulfide bond (Fig. 8A) (38). In the absence of the disulfide bond, therefore, generation of the zinc-binding site in E. coli SodC may be hampered by significant fluctuations of the relatively long S-S subloop.…”

Section: Discussionmentioning

confidence: 99%

A Primary Role for Disulfide Formation in the Productive Folding of Prokaryotic Cu,Zn-superoxide Dismutase

Sakurai

Anzai

Furukawa

2014

Journal of Biological Chemistry

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Unique structural features of the monomeric Cu,Zn superoxide dismutase from Escherichia coli, revealed by X-ray crystallography

Cited by 81 publications

References 49 publications

A prokaryotic superoxide dismutase paralog lacking two Cu ligands: From largely unstructured in solution to ordered in the crystal

A prokaryotic superoxide dismutase paralog lacking two Cu ligands: From largely unstructured in solution to ordered in the crystal

Role of the electrostatic loop charged residues in Cu, Zn superoxide dismutase

A Primary Role for Disulfide Formation in the Productive Folding of Prokaryotic Cu,Zn-superoxide Dismutase

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