Electrostatic recognition between superoxide and copper, zinc superoxide dismutase

Abstract

Electrostatic forces have been implicated in a variety of biologically important molecular interactions including drug orientation by DNA, protein folding and assembly, substrate binding and catalysis and macromolecular complementarity with inhibitors, drugs and hormones. To examine enzyme-substrate interactions in copper, zinc superoxide dismutase (SOD), we developed a method for the visualization and analysis of an enzyme's three-dimensional electrostatic vector field that allows the contributions of specific residues to be identified. We report here that the arrangement of electrostatic charges in SOD promotes productive enzyme-substrate interaction through substrate guidance and charge complementarity: sequence-conserved residues create an extensive electrostatic field that directs the negatively charged superoxide (O-2) substrate to the highly positive catalytic binding site at the bottom of the active-site channel. Dissection of the electrostatic potential gradient indicated the relative contributions of individual charged residues: Lys 134 and Glu 131 seem to have important roles in directing the long-range approach of O-2, while Arg 141 has local orienting effects. The reported methods of analysis may have general application for the elucidation of intermolecular recognition processes.

Autoxidation of Ubiquinol-6 Is Independent of Superoxide Dismutase

“…Alternatively, it has been postulated that superoxide dismutase [EC 1.15.1.1] (SOD) may protect quinols from autoxidation by functioning as a superoxide-semiquinone oxidoreductase (eq 8) (Cadenas et al, 1988(Cadenas et al, , 1992. However, the direct reduction of QH • by SOD seems unlikely, since the crystal structure of SOD indicates the presence of a narrow cleft in the active site channel which restricts access to superoxide and other ions and competitive inhibitors smaller than 4 Å Getzoff et al, 1983).…”

The free radical‐generating function of a familial amyotrophic lateral sclerosis‐associated D90A Cu,Zn‐superoxide dismutase mutant

“…3B, curves a and c) show a large decrease in ABTS +" adduct concentrations when azide is present. X-ray crystallographic studies [16] revealed that Arg-14 l inside and Lys-120 and Lys-134 at the active channel of the Cu,Zn-SOD may be responsible for the favorable electrostatic guidance of the anionic substrate 02-" to the active site to yield the superoxide dismutation. Other small anions, such as cyanide, azide, halides and phosphate, are also known to have easy access inside the channel and to bind to and the human Cu,Zn-SOD.…”

Protein engineering. The design, synthesis and characterization of factitious proteins

“…Both the genes for a soluble form of cytochrome b5, deleted for the membrane anchor segment, and also that for the complete protein were constructed. The soluble microbial product was shown to possess the properties expected for natural cytochrome b5, and the first round of mutagenesis experiments have been directed at the roles played by (Tainer et al, 1982Getzoff et al, 1983). In addition to its importance as a biological defence mechanism against the superoxide anion (02-), superoxide dismutase has a number of structural and mechanistic features which lend themselves to study by directed mutagenesis .…”