Lipid-destabilizing properties of the hydrophobic helices H8 and H9 from colicin E1

Colicins are toxic proteins produced by Escherichia coli that must cross the membrane to exert their activity. The lipid insertion of their pf domain is linked to a conformational change which enables the penetration of a hydrophobic hairpin. They provide useful models to more generally study insertion of proteins, channel formation and protein translocation in and across membranes. In this paper, we study the lipid-destabilizing properties of helices H8 and H9 forming the hydrophobic hairpin of colicin E1. Modelling analysis suggests that those fragments behave like tilted peptides. The latter are characterized by an asymmetric distribution of their hydrophobic residues when helical. They are able to interact with a hydrophobic/hydrophilic interface (such as a lipid membrane) and to destabilize the organized system into which they insert. Fluorescence techniques using labelled liposomes clearly show that H9, and H8 to a lesser extent, destabilize lipid particles, by inducing fusion and leakage. AFM assays clearly indicate that H8 and especially H9 induce membrane fragilization. Holes in the membrane are even observed in the presence of H9. This behaviour is close to what is seen with viral fusion peptides. Those results suggest that the peptides could be involved in the toroidal pore formation of colicin E1, notably by disturbing the lipids and facilitating the insertion of the other, more hydrophilic, helices that will form the pore. Since tilted, lipid-destabilizing fragments are also common to membrane proteins and to signal sequences, we suggest that tilted peptides should have an ubiquitous role in the mechanism of insertion of proteins into membranes.     

GM2/GM3 controls the organizational status of CD82/Met microdomains: further studies in GM2/GM3 complexation

Glycoconjugate Journal - At cell surface gangliosides might associate with signal transducers proteins, grown factor receptors, integrins, small G-proteins and tetraspanins establishing...     

Occurrence of south- and north-seeking multicellular magnetotactic prokaryotes in a coastal lagoon in the South Hemisphere

International Microbiology - Magnetotactic bacteria (MTB) response to the magnetic field can be classified into north-seeking (NS) and south-seeking (SS), which usually depends on their inhabiting...     

Complementary biophysical tools to investigate lipid specificity in the interaction between bioactive molecules and the plasma membrane: A review

Plasma membranes are complex entities common to all living cells. The basic principle of their organization appears very simple, but they are actually of high complexity and represent very dynamic structures. The interactions between bioactive molecules and lipids are important for numerous processes, from drug bioavailability to viral fusion. The cell membrane is a carefully balanced environment and any change inflicted upon its structure by a bioactive molecule must be considered in conjunction with the overall effect that this may have on the function and integrity of the membrane. Conceptually, understanding the molecular mechanisms by which bioactive molecules interact with cell membranes is of fundamental importance.     

Human recombinant thiamine triphosphatase: purification, secondary structure and catalytic properties

Thiamine triphosphate (ThTP) is found in most living organisms and it may act as a phosphate donor for protein phosphorylation. We have recently cloned the cDNA coding for a highly specific mammalian 25 kDa thiamine triphosphatase (ThTPase; EC 3.6.1.28). As the enzyme has a high catalytic efficiency and no sequence homology with known phosphohydrolases, it was worth investigating its structure and catalytic properties. For this purpose, we expressed the untagged recombinant human ThTPase (hThTPase) in E. coli, produced the protein on a large scale and purified it to homogeneity. Its kinetic properties were similar to those of the genuine human enzyme, indicating that the recombinant hThTPase is completely functional. Mg2+ ions were required for activity and Ca2+ inhibited the enzyme by competition with Mg2+. With ATP as substrate, the catalytic efficiency was 10(-4)-fold lower than with ThTP, confirming the nearly absolute specificity of the 25 kDa ThTPase for ThTP. The activity was maximum at pH 8.5 and very low at pH 6.0. Zn2+ ions were inhibitory at micromolar concentrations at pH 8.0 but activated at pH 6.0. Kinetic analysis suggests an activator site for Mg2+ and a separate regulatory site for Zn2+. The effects of group-specific reagents such as Woodward's reagent K and diethylpyrocarbonate suggest that at least one carboxyl group in the active site is essential for catalysis, while a positively charged amino group may be involved in substrate binding. The secondary structure of the enzyme, as determined by Fourier-transform infrared spectroscopy, was predominantly beta-sheet and alpha-helix.     

Oxidation of dibenzothiophene (DBT) by Serratia marcescens UCP 1549 formed biphenyl as final product

ABSTRACT: BACKGROUND: The desulphurization of dibenzothiophene (DBT), a recalcitrant thiophenic fossil fuel component by Serratia marcescens (UCP 1549) in order for reducing the sulphur content was investigated. The study was carried out establishing the growth profile using Luria Bertani medium to different concentrations of DBT during 120hours at 28oC, and orbital shaker at 150rpm. RESULTS: The results indicated that concentrations of DBT 0.5, 1.0 and 2.0 mM do not affected the growth of the bacterium. The DBT showed similar Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MCB) (3.68 mM). The desulphurization of DBT by S. marcescens was used with 96 hours of growth on 2mM of DBT, and was determined by gas chromatography (GC) and GC-mass spectrometry. In order to study the desulphurization process by S. marcescens was observed the presence of a sulfur-free product at 16 hours of cultivation The results show that S. marcescens oxidizes DBT to its corresponding DBT-5 oxide and then to DBT-sulfone, without the formation of any biphenyl. CONCLUSIONS: The data suggests the use of metabolic pathway "4S" by S. marcescens (UCP 1549) and formed biphenyl. The microbial desulphurization process by S. Serratia can be suggest significant reducing sulphur content in DBT, and showed promising potential for reduction of the sulfur content in diesel oil.     

Remaining genetic diversity in Brazilian Merganser (Mergus octosetaceus)

The Brazilian Merganser is a very rare and threatened species that nowadays inhabits only a few protected areas and their surroundings in the Brazilian territory. In order to estimate the remaining genetic diversity and population structure in this species, two mitochondrial genes were sequenced in 39 individuals belonging to two populations and in one individual collected in Argentina in 1950. We found a highly significant divergence between two major remaining populations of Mergus octosetaceus, which suggests a historical population structure in this species. Furthermore, two deeply divergent lineages were found in a single location, which could due to current or historical secondary contact. Based on the available genetic data, we point out future directions which would contribute to design strategies for conservation and management of this threatened species.     

Characterization of the interactions between fluoroquinolone antibiotics and lipids: a multitechnique approach

Probing drug/lipid interactions at the molecular level represents an important challenge in pharmaceutical research and membrane biophysics. Previous studies showed differences in accumulation and intracellular activity between two fluoroquinolones, ciprofloxacin and moxifloxacin, that may actually result from their differential susceptibility to efflux by the ciprofloxacin transporter. In view of the critical role of lipids for the drug cellular uptake and differences observed for the two closely related fluoroquinolones, we investigated the interactions of these two antibiotics with lipids, using an array of complementary techniques. Moxifloxacin induced, to a greater extent than ciprofloxacin, an erosion of the DPPC domains in the DOPC fluid phase (atomic force microscopy) and a shift of the surface pressure-area isotherms of DOPC/DPPC/fluoroquinolone monolayer toward lower area per molecule (Langmuir studies). These effects are related to a lower propensity of moxifloxacin to be released from lipid to aqueous phase (determined by phase transfer studies and conformational analysis) and a marked decrease of all-trans conformation of acyl-lipid chains of DPPC (determined by ATR-FTIR) without increase of lipid disorder and change in the tilt between the normal and the germanium surface (also determined by ATR-FTIR). All together, differences of ciprofloxacin as compared to moxifloxacin in their interactions with lipids could explain differences in their cellular accumulation and susceptibility to efflux transporters.     

The minimal fusion peptide of simian immunodeficiency virus corresponds to the 11 first residues of gp32.

We had previously predicted successfully the minimal fusion peptides (FPs) of the human immunodeficiency virus 1 (HIV-1) gp41 and the bovine leukemia virus (BLV) gp30 using an original approach based on the obliquity/fusogenicity relationship of tilted peptides. In this paper, we have used the same method to predict the shortest FP capable of inducing optimal fusion in vitro of the simian immunodeficiency virus (SIV) mac isolate and of other SIVs and human immunodeficiency virus (HIV-2) isolates. In each case, the 11-residue-long peptide was predicted as the minimal FP. For the SIV mac isolate, liposome lipid-mixing and leakage assays confirmed that this peptide is the shortest peptide inducing optimal fusion in vitro, being therefore the minimal FP. These results are another piece of evidence that the tilted properties of FPs are important for the fusion process and that our method can be used to predict the minimal FPs of other viruses.