More Than One Way to Skin a Cat

The first structural pictures of a nickel-containing enzyme that destroys harmful superoxide radicals (O₂^–) recently were reported. In addition to revealing a novel nickel catalytic center, the structures demonstrate the remarkable number of methods nature has devised to destroy these ubiquitous reactive oxygen species.

To guard against the dangers of highly reactive superoxide radicals, which are a by-product of normal metabolism, organisms from bacteria to humans rely on enzymes called superoxide dismutases (SODs). These enzymes convert superoxide into peroxide and water; other cellular enzymes then break down the peroxide into water and oxygen.

But nature has shown that there's more than one way to make an SOD. Human cells make both a copper-and-zinc SOD and a manganese version that has a completely different amino acid sequence and structure. Bacteria and some plants make an iron SOD similar to manganese SOD. Most recently, scientists discovered a novel nickel-containing SOD in Streptomyces soil bacteria. Now, the first pictures of the nickel enzyme show that it looks nothing like previously characterized SODs, illustrating just how ingenious and flexible nature really is.

"Nature has evolved three different folds capable of catalyzing the same reaction," says crystallographer Elizabeth D. Getzoff of Scripps Research Institute. "That nature has figured out how to use these different metals to do the same chemistry is really quite remarkable," she adds.

Getzoff, postdoc David P. Barondeau, and coworkers reported a 1.30-Å-resolution structure of the nickel SOD found in the well-studied soil bacterium.

Streptomyces coelicolor [Biochemistry, 43, 8038 (2004)]. Their report closely followed a South Korean and Italian team's 1.6- and 1.68-Å structures of nickel SOD from

Streptomyces seoulensis, a Korean soil bacterium [Proc. Natl. Acad. Sci. USA, 101, 8569 (2004)]. The team was led by biophysics professor Sa-Ouk Kang of Seoul National University in South Korea and crystallographer Kristina Djinovic Carugo of the Synchrotron Light Laboratory in Trieste, Italy.

In both structures, six identical, mostly a-helical subunits cluster to form a hollow sphere. Each subunit has an unstructured, nine-residue tail at its N-terminus that folds up into a hook when it binds a single nickel ion.

"By itself, the nickel hook provides almost all interactions critical for metal binding and catalysis," contributing not just all of the nickel ligands but also residues that help stabilize and guide superoxide through the protein to the nickel centers that rim the central cavity, Barondeau points out. He and Getzoff are currently testing whether the nickel hook can catalyze the SOD reaction on its own.

Unlike the other metal ions nature uses in SODs, Ni²⁺ normally cannot access the redox potentials required to convert superoxide into peroxide and water. To carry out the SOD reaction, nickel SOD must somehow stabilize the Ni³⁺/Ni²⁺ couple. In fact, the tricks the enzyme uses--chelating nickel via two thiolates and two nitrogens and expanding nickel's coordination sphere--are well known to nickel coordination chemists, notes inorganic chemist Marcetta Y. Darensbourg of Texas A&M University.

 THE ENZYME coordinates the reduced Ni²⁺ center via two cysteine thiolates, a backbone nitrogen atom, and the N-terminus of the protein in a square-planar geometry. When a histidine ring nitrogen swings in to become the axial Ni ligand, the resulting five-coordinate, square pyramidal site favors oxidation to Ni³⁺. This small change "tunes the reduction potential for superoxide dismutation," Getzoff says.

What's most remarkable about the structure is the nickel hook domain, according to Scripps crystallographer John A. Tainer, a coauthor of the Biochemistry paper. Most metalloenzymes, he points out, use residues from different parts of their amino acid sequence to make a preorganized active site for a catalytic metal atom. In contrast, nickel SOD's unstructured N-terminal peptide wraps around a nickel ion to create a catalytic site. This "suggests ways to build low-molecular-weight, catalytic metal-ion compounds and to design novel catalytic metal-ion sites in proteins," Tainer says.

Tainer also suggests that the peptide could be used as an alternative to the popular nickel-binding "His₆" tags used to purify recombinant proteins or as a spectroscopic tag. Crystallographers could also find it useful to make heavy-atom derivatives of their protein targets for X-ray structure determination, Getzoff adds.