New insights into the co-evolution of cytochrome c reductase and the mitochondrial processing peptidase

The mitochondrial processing peptidase (MPP) is a heterodimeric enzyme that forms part of the cytochrome c reductase complex from higher plants. Mitochondria from mammals and yeast contain two homologous enzymes: (i) an active MPP within the mitochondrial matrix and (ii) an inactive MPP within the cytochrome c reductase complex. To elucidate the evolution of MPP, the cytochrome c reductase complexes from lower plants were isolated and tested for processing activity. Mitochondria were prepared from the staghorn fern Platycerium bifurcatum, from the horsetail Equisetum arvense, and from the colorless algae Polytomella, and cytochrome c reductase complexes were purified by a micro-isolation procedure based on Blue-native polyacrylamide gel electrophoresis and electroelution. This is the first report on the subunit composition of a respiratory enzyme complex from a fern or a horsetail. The cytochrome c reductase complexes from P. bifurcatum and E. arvense are shown to efficiently process mitochondrial precursor proteins, whereas the enzyme complex from Polytomella lacks proteolytic activity. An evolutionary model is suggested that assumes a correlation between the presence of an active MPP within the cytochrome c reductase complex and the occurrence of chloroplasts

Impact of anion polarizability on ion pairing in microhydrated salt clusters

Despite longstanding interest in the mechanism of salt dissolution in aqueous media, a molecular level understanding remains incomplete. Here, cryogenic ion trap vibrational action spectroscopy is combined with electronic structure calculations to track salt hydration in a gas phase model system one water molecule at a time. The infrared photodissociation spectra of microhydrated lithium dihalide anions [LiXX′(H2O)n]- (XX′ = I2, ClI and Cl2; n = 1–3) in the OH stretching region (3800–2800 cm-1) provide a detailed picture of how anion polarizability influences the competition among ion–ion, ion–water and water–water interactions. While exclusively contact ion pairs are observed for n = 1, the formation of solvent-shared ion pairs, identified by markedly red-shifted OH stretching bands (-1), originating from the bridging water molecules, is favored already for n = 2. For n = 3, Li+ reaches its maximum coordination number of four only in [LiI2(H2O)3]-, in accordance with the hard and soft Lewis acid and base principle. Water–water hydrogen bond formation leads to a different solvent-shared ion pair motif in [LiI2(H2O)3]- and network formation even restabilizes the contact ion pair motif in [LiCl2(H2O)3]-. Structural assignments are exclusively possible after the consideration of anharmonic effects. Molecular dynamics simulations confirm that the significance of large amplitude motion (of the water molecules) increases with increasing anion polarizability and that needs to be considered already at cryogenic temperatures

HIV-1 subtype C Nef-mediated SERINC5 down-regulation significantly contributes to overall Nef activity

BACKGROUND: Nef performs multiple cellular activities that enhance HIV-1 pathogenesis. The role of Nef-mediated down-regulation of the host restriction factor SERINC5 in HIV-1 pathogenesis is not well-defined. We aimed to investigate if SERINC5 down-regulation activity contributes to HIV-1 subtype C disease progression, to assess the relative contribution of this activity to overall Nef function, and to identify amino acids required for optimal activity. We measured the SERINC5 down-regulation activity of 106 subtype C Nef clones, isolated from individuals in early infection, for which the Nef activities of CD4 and HLA-I down-regulation as well as alteration of TCR signalling were previously measured. The relationship between SERINC5 down-regulation and markers of disease progression, and the relative contribution of SERINC5 down-regulation to a Nef fitness model-derived E value (a proxy for overall Nef fitness in vivo), were assessed. RESULTS: No overall relationship was found between SERINC5 down-regulation and viral load set point (p = 0.28) or rate of CD4+ T cell decline (p = 0.45). CD4 down-regulation (p = 0.02) and SERINC5 down-regulation (p = 0.003) were significant determinants of E values in univariate analyses, with the greatest relative contribution for SERINC5 down-regulation, and only SERINC5 down-regulation remained significant in the multivariate analysis (p = 0.003). Using a codon-by-codon analysis, several amino acids were significantly associated with increased (10I, 11V, 38D, 51T, 65D, 101V, 188H and, 191H) or decreased (10K, 38E, 65E, 135F, 173T, 176T and, 191R) SERINC5 down-regulation activity. Site-directed mutagenesis experiments of selected mutants confirmed a substantial reduction in SERINC5 down-regulation activity associated with the mutation 173T, while mutations 10K, 135F, and 176T were associated with more modest reductions in activity that were not statistically significant. CONCLUSIONS: These results suggest that SERINC5 down-regulation is a significant contributor to overall Nef function and identify potential genetic determinants of this Nef function that may have relevance for vaccines or therapeutics