A broad distribution of the alternative oxidase in microsporidian parasites

Microsporidia are a group of obligate intracellular parasitic eukaryotes that were considered to be amitochondriate until the recent discovery of highly reduced mitochondrial organelles called mitosomes. Analysis of the complete genome of Encephalitozoon cuniculi revealed a highly reduced set of proteins in the organelle, mostly related to the assembly of ironsulphur clusters. Oxidative phosphorylation and the Krebs cycle proteins were absent, in keeping with the notion that the microsporidia and their mitosomes are anaerobic, as is the case for other mitosome bearing eukaryotes, such as Giardia. Here we provide evidence opening the possibility that mitosomes in a number of microsporidian lineages are not completely anaerobic. Specifically, we have identified and characterized a gene encoding the alternative oxidase (AOX), a typically mitochondrial terminal oxidase in eukaryotes, in the genomes of several distantly related microsporidian species, even though this gene is absent from the complete genome of E. cuniculi. In order to confirm that these genes encode functional proteins, AOX genes from both A. locustae and T. hominis were over-expressed in E. coli and AOX activity measured spectrophotometrically using ubiquinol-1 (UQ-1) as substrate. Both A. locustae and T. hominis AOX proteins reduced UQ-1 in a cyanide and antimycin-resistant manner that was sensitive to ascofuranone, a potent inhibitor of the trypanosomal AOX. The physiological role of AOX microsporidia may be to reoxidise reducing equivalents produced by glycolysis, in a manner comparable to that observed in trypanosome

MODELING VANADIUM BROMOPEROXIDASE - SYNTHESIS, STRUCTURE, AND SPECTRAL PROPERTIES OF VANADIUM(IV) COMPLEXES WITH COORDINATED IMIDAZOLE

A series of vanadium(IV)-SALIMH complexes [HSALIMH = 4-(2-(salicylideneamino)ethyl)imidazole] have been synthesized and characterized by X-ray crystallography, electron paramagnetic resonance, infrared, and visible spectroscopy, and mass spectrometry. The compounds VO(SALIMH)ACAC??MeOH (1), VO(SALIMH)SAL??MeOH (SAL is the anion of salicyl-aldehyde) (2), and VO(SALIMH)2??3EtOH (3) represent the first fully characterized vanadium(IV) compounds with imidazole in their coordination sphere. Compounds 1-3 react reversibly with acid to form complexes in which either the bound imidazole has been protonated (1??? and 3???) or the bidentate ligand has been protonated and subsequently displaced by solvent (4). Addition of excess acid yields the solvated vanadyl ion VO(SOL)52+ (5) (SOL = solvent). Protonation and subsequent ligand displacement by solvent induce an increase in the EPR hyperfine coupling constants A??? and A???. The implications of this work toward understanding the reduced catalytically inactive form of vanadium bromoperoxidase are discussed. X-ray parameters are as follows: 1, C18H23N3O5V1, 412.34 g/mol, crystal system, orthorhombic (Pbca), a = 20.673 (4) A, b = 21.335 (4) A, c = 9.078 (1) A, V = 4404 (1) A3, Z = 8, 2629 data collected with 5?? < 2?? < 45??, 1807 data collected with Fo > 6??(F), R = 0.039, Rw = 0.038; 2, C20H21N3O5V1, 434.34 g/mol, crystal system, orthorhombic (Pbca), a = 12.336 (5) A, b = 14.928 (6) A, c = 22.03 (1) A, V = 4067 (3) A3, Z = 8, 3588 data collected with 5?? < 2?? < 50??, 2723 data collected with Fo > 2.0??(F), R = 0.0799, Rw = 0.0479; 3, C30H42N6O6V1, 633.64 g/mol, crystal system, monoclinic (P21/c), a = 13.763 (7) A, b = 17.589 (4) A, c = 14.701 (2) A, ?? = 108.97 (2)??, V= 3365 (1) A3, Z = 4,4398 data collected with 5?? < 2?? < 45??, 3059 data collected with Fo > 5??(F) R = 0.0697, Rw = 0.0949.close1059

The genome of the obligate intracellular parasite Trachipleistophora hominis : new insights into microsporidian genome dynamics and reductive evolution

The dynamics of reductive genome evolution for eukaryotes living inside other eukaryotic cells are poorly understood compared to well-studied model systems involving obligate intracellular bacteria. Here we present 8.5 Mb of sequence from the genome of the microsporidian Trachipleistophora hominis, isolated from an HIV/AIDS patient, which is an outgroup to the smaller compacted-genome species that primarily inform ideas of evolutionary mode for these enormously successful obligate intracellular parasites. Our data provide detailed information on the gene content, genome architecture and intergenic regions of a larger microsporidian genome, while comparative analyses allowed us to infer genomic features and metabolism of the common ancestor of the species investigated. Gene length reduction and massive loss of metabolic capacity in the common ancestor was accompanied by the evolution of novel microsporidian-specific protein families, whose conservation among microsporidians, against a background of reductive evolution, suggests they may have important functions in their parasitic lifestyle. The ancestor had already lost many metabolic pathways but retained glycolysis and the pentose phosphate pathway to provide cytosolic ATP and reduced coenzymes, and it had a minimal mitochondrion (mitosome) making Fe-S clusters but not ATP. It possessed bacterial-like nucleotide transport proteins as a key innovation for stealing host-generated ATP, the machinery for RNAi, key elements of the early secretory pathway, canonical eukaryotic as well as microsporidian-specific regulatory elements, a diversity of repetitive and transposable elements, and relatively low average gene density. Microsporidian genome evolution thus appears to have proceeded in at least two major steps: an ancestral remodelling of the proteome upon transition to intracellular parasitism that involved reduction but also selective expansion, followed by a secondary compaction of genome architecture in some, but not all, lineages.Publisher PDFPeer reviewe