The active site of the plant alternative oxidase: structural and mechanistic considerations

Abstract

In the present review we seek to provide an up‐to‐date view on the molecular nature of the active site of the plant alternative oxidase which has been postulated to comprise of a binuclear iron centre. A three‐dimensional model of the catalytic centre of the oxidase is presented which is based on the active site structure of the free radical component of ribonucleotide reductase and methane monooxygenase. The model indicates that a highly conserved carboxylate (Glu‐270) occupies a central position within the proposed di‐iron centre as it co‐ordinates both iron atoms. The expression of an alternative oxidase protein in Schizosaccharomyces pombe in which Glu‐270 was mutated to asparagine yields an inactive protein. The implications of this in relation to the structural model of the active site of the oxidase suggests that Glu‐270 is essential for catalytic alternative oxidase activity. A kinetic mechanism is suggested which accounts for the full reduction of dioxygen to water catalysed by a single di‐iron centre. © 2000 Society of Chemical Industry

Plant Mitochondria: From Genome to Function

“…This bundle brought D288 of AtAOXla and two of the three E-X-X-H motifs [E275-X-X-H278 and E324-X-X-H327] into a position that would allow the binding of two iron atoms by five of the six ligands found in the typical di-iron carboxylate proteins. The lack of a sixth amino-acid ligand was justified by analogy to other members of the di-iron carboxylate family that only have five ligands (Siedow et al, 1995) and has been postulated to introduce flexibility to the AOX active site (Affourtit et al, 2000), as is common for di-iron carboxylate proteins (Nordlund and Eklund, 1995;Berthold and Stenmark, 2003).…”

“…Several catalytic mechanisms for AOX have been proposed and the possible involvement of several absolutely, or nearly so, conserved residues of the Dox sequence family as possible radical intermediates has been highlighted (Moore et al, Affourtit et al, 2000Affourtit et al, , 2002Berthold eta!., 2000Berthold eta!., , 2002Albury et al, 2002). The position of the absolutely conserved Y280 (Fig.…”

Exploring the molecular nature of alternative oxidase regulation and catalysis

“…These weaknesses, together with the notion that one of the iron‐co‐ordinating histidines (H273) is not fully conserved throughout the by then numerous newly emerged alternative oxidase sequences, provoked these authors to revise the model proposed by Siedow and colleagues [29]. Using an until recently [30] ignored but indeed fully conserved glutamate‐histidine motif (E217‐X‐X‐H220), a structure was proposed in which there is a greater degree of resemblance to other di‐iron enzymes. Arguably the most distinct feature of the revised model is that the alternative oxidase protein is no longer predicted to be trans‐membranal, but rather to be peripherally associated with the matrix side of the inner membrane (Fig.…”

“…The steadily increasing relative wealth of information on the structure of reaction intermediates of other di‐iron enzymes (see e.g. [27]) led to the proposal of two alternative catalytic models that both address this ‘overload’ aspect [13,30]. Although similar in nature, these models differ as to the exact sequence in which oxygen and quinol bind to the enzyme.…”

“…Although similar in nature, these models differ as to the exact sequence in which oxygen and quinol bind to the enzyme. Furthermore, the reaction mechanism suggested by Affourtit et al [30] involves a high‐valent diferryl intermediate, whereas Berthold and colleagues [13] questioned the catalytic necessity of a compound with such strong oxidising potential and hence did not include it in their model.…”