An evolutionary path to altered cofactor specificity in a metalloenzyme

Abstract

Almost half of all enzymes utilize a metal cofactor. However, the features that dictate the metal utilized by metalloenzymes are poorly understood, limiting our ability to manipulate these enzymes for industrial and health-associated applications. The ubiquitous iron/manganese superoxide dismutase (SOD) family exemplifies this deficit, as the specific metal used by any family member cannot be predicted. Biochemical, structural and paramagnetic analysis of two evolutionarily related SODs with different metal specificity produced by the pathogenic bacterium Staphylococcus aureus identifies two positions that control metal specificity. These residues make no direct contacts with the metal-coordinating ligands but control the metal’s redox properties, demonstrating that subtle architectural changes can dramatically alter metal utilization. Introducing these mutations into S. aureus alters the ability of the bacterium to resist superoxide stress when metal starved by the host, revealing that small changes in metal-dependent activity can drive the evolution of metalloenzymes with new cofactor specificity.

Substitution of histidine 30 by asparagine in manganese superoxide dismutase alters biophysical properties and supports proliferation in a K562 leukemia cell line

“…The shape and width are the consequence of the large GHz Mn(II) zero-field interaction of the penta-coordinate Mn(II) center in SODs. This larger zero-field interaction is a characteristic of Mn(II)SODs and its specific magnitude highly correlates with the Mn or Fe metal specificity (Barwinska-Sendra et al 2020 ). Upon addition of the azide, a narrower six-line component (from 10.13 to 10.20) was observed.…”

Interchangeable utilization of metals: New perspectives on the impacts of metal ions employed in ancient and extant biomolecules

“…These structural similarities suggest that cambialism is not provided by the inner sphere coordination geometry but relies on differences in the secondary coordination sphere. Mutational analysis has identified two amino acids present in positions 159 and 160 (Figure 4B) that vary between SodA (possesses Gly159 and Leu160) and SodM (Leu159, Phe160) that make no direct contact but are in close proximity (<10Å) to a metal co-factor (147). Significantly, swapping these amino acids between SodA and SodM did not affect active center structures but enabled cambialistic properties to SodA (147).…”

“…Mutational analysis has identified two amino acids present in positions 159 and 160 (Figure 4B) that vary between SodA (possesses Gly159 and Leu160) and SodM (Leu159, Phe160) that make no direct contact but are in close proximity (<10Å) to a metal co-factor (147). Significantly, swapping these amino acids between SodA and SodM did not affect active center structures but enabled cambialistic properties to SodA (147). It was proposed that amino acid side chains in positions 159 and 160 are responsible for changes in the reduction potential of the metal ions, likely underlying the mechanism of catalysis governed by Mn and Fe ions (147).…”

Coping with oxidative stress in extreme environments: the distinctive roles played by Acinetobacter sp. Ver3 superoxide dismutases

“…2A displays a multiple sequence alignment of AV3 SodB and three structurally characterized related enzymes: the FeSOD (PDB code 1ISA) and MnSOD (PDB code 1VEW) from E. coli (60 % and 44 % amino acid identity, respectively) and the cambialistic SOD (PDB code 1QNN) from P. gingivalis (50 % identity). It shows that AV3 SodB harbours motifs that are characteristic of FeSODs, such as the motifs AAQ and DVEWHAYY involved in catalysis 39 . A phylogenetic tree built for AV3 SodB and diverse FeSODs, MnSODs as well as cambialistic Fe/MnSODs (here collectively referred to as Fe-MnSODs) of known structure consisted of three different clades, with archaeal and bacterial FeSODs grouped in two different clades whereas bacterial MnSODs constituted a third one (Fig.…”