Abstract
The alternative oxidase (AOX) protein is present in plants, fungi, protozoa and some invertebrates. It is involved in the mitochondrial respiratory chain, providing an alternative route for the transport of electrons, leading to the reduction of oxygen to form water. The present study aimed to characterize the family of AOX genes in mandarin (Citrus clementina) and sweet orange (Citrus sinensis) at nucleotide and protein levels, including promoter analysis, phylogenetic analysis and C. sinensis gene expression. This study also aimed to do the homology modeling of one AOX isoform (CcAOXd). Moreover, the molecular docking of the CcAOXd protein with the ubiquinone (UQ) was performed. Four AOX genes were identified in each citrus species. These genes have an open reading frame (ORF) ranging from 852 bp to 1150 bp and a number of exons ranging from 4 to 9. The 1500 bp-upstream region of each AOX gene contained regulatory cis-elements related to internal and external response factors. CsAOX genes showed a differential expression in citrus tissues. All AOX proteins were predicted to be located in mitochondria. They contained the conserved motifs LET, NERMHL, LEEEA and RADE-H as well as several putative post-translational modification sites. The CcAOXd protein was modeled by homology to the AOX of Trypanosona brucei (45% of identity). The 3-D structure of CcAOXd showed the presence of two hydrophobic helices that could be involved in the anchoring of the protein in the inner mitochondrial membrane. The active site of the protein is located in a hydrophobic environment deep inside the AOX structure and contains a diiron center. The molecular docking of CcAOXd with UQ showed that the binding site is a recessed pocket formed by the helices and submerged in the membrane. These data are important for future functional studies of citrus AOX genes and/or proteins, as well as for biotechnological approaches leading to AOX inhibition using UQ homologs.
Highlights
The term oxidase refers to any enzyme that catalyzes the oxidation–reduction reaction involving molecular oxygen as an electron acceptor
The number of alternative oxidase (AOX) genes found in C. clementina and C. sinensis is small, which is similar to what has been observed in other species such as Arabidopsis thaliana, whose AOX family is represented by five genes [7]; Glycine max [32], Oryza sativa [33] and Zea mays [34], each represented by three genes; and Nicotiana tabacum [2], Triticum aestivum [35] and Hypericum perforatum [36], each represented by two genes
Phylogenetic analysis of the AOX citrus and A. thaliana sequences showed that the CcAOXb and CsAOXb were closed to the AtAOX1D sequence while CcAOXd and CsAOXd were closed to AtAOX2 (Fig 4)
Summary
The term oxidase refers to any enzyme that catalyzes the oxidation–reduction reaction involving molecular oxygen as an electron acceptor. The main electron transport route in eukaryotes passes through the complex IV (known as cyanide-sensitive cytochrome oxidase) of the respiratory chain, but in some organisms the electron transport route goes through the AOX protein (known as cyanide-insensitive and hydroxamic acid-sensitive terminal oxidase). Both routes lead to the transportation of electrons and the reduction of oxygen to form water [1, 2]. The AOX catalyzes the four-electron oxidation of ubiquinol (reduced form of ubiquinone [UQ]) by oxygen, and the energy of ubiquinol oxidation by oxygen is released as heat [3,4,5]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
