The Q(y) absorption spectrum of the light-harvesting complex II as determined by structure-based analysis of chlorophyll macrocycle deformations

The absorption spectrum of the main antenna complex of photosystem II, LHCII, has been modeled using, as starting points, the chlorophyll (chl) atomic coordinates as obtained by the LHCII crystal analysis [Liu, Z., Yan, H., Wang, K., Kuang, T., Zhang, J., Gui, L., An, X., and Chang, W. (2004) Nature 428, 287-292] of three different trimers. The chl site Q(y) transition energies have been obtained in terms of the chl macrocycle deformations influencing the energy level of the chl frontier orbitals. Using these chl site transition energy values and the entire set of interaction energies, calculated in the ideal dipole approximation, the complete Hamiltonians for the three LHCII trimers have been written and the full set of 42 eigenstates of each LHCII trimer have been calculated. With the 42 transition energies and transition dipole strengths, either unperturbed or associated to the eigenstates, the LHCII Q(y) absorption spectrum has been calculated using a chl absorption band shape. These calculations have been performed both in vacuo and in the presence of a medium. Despite the number of approximations, a good correlation with the measured absorption spectrum of a LHCII preparation is obtained. This analysis shows that, although a substantial C3 symmetry of the LHCII trimer in terms of both chl-chl distances and interaction energies is present, a marked variation among monomer subsets of site transition energies is estimated. This leads to a C3 symmetry breaking in the unperturbed chl site transition energies set and, consequently, in the trimer eigenstates. It is also concluded that interactions among chlorophylls do not significantly modify the light absorption role of LHCII in plant leaves.     

Dynamics-stability relationships in apo- and holomyoglobin: a combined neutron scattering and molecular dynamics simulations study

The removal of the heme group from myoglobin (Mb) results in a destabilization of the protein structure. The dynamic basis of the destabilization was followed by comparative measurements on holo- (holo-Mb) and apomyoglobin (apo-Mb). Mean-squared displacements (MSD) and protein resilience on the picosecond-to-nanosecond timescale were measured by elastic incoherent neutron scattering. Differences in thermodynamic parameters, MSD, and resilience were observed for both proteins. The resilience of holo-Mb was significantly lower than that of apo-Mb, indicating entropic stabilization by a higher degree of conformational sampling in the heme-bound folded protein. Molecular dynamics simulations provided site-specific information. Averaged over the whole structure, the molecular dynamics simulations yielded similar MSD and resilience values for the two proteins. The mobility of residues around the heme group in holo-Mb showed a smaller MSD and higher resilience compared to the same residue group in apo-Mb. It is of interest that in holo-Mb, higher MSD values are observed for the residues outside the heme pocket, indicating an entropic contribution to protein stabilization by heme binding, which is in agreement with experimental results.     

Over-expression in Escherichia coli, purification and reconstitution in liposomes of the third member of the OCTN sub-family: the mouse carnitine transporter OCTN3

pET-21a(+)-mOCTN3-6His was constructed and used for over-expression in Escherichia coli Rosetta(DE3)pLysS. After IPTG induction a protein with apparent molecular mass of 53 kDa was collected in the insoluble fraction of the cell lysate and purified by Ni(2+)-chelating chromatography with a yield of 2mg/l of cell culture. The over-expressed protein was identified with mOCTN3 by anti-His antibody and reconstitution in liposomes. mOCTN3 required peculiar conditions for optimal expression and reconstitution in liposomes. The protein catalyzed a time dependent [(3)H]carnitine uptake which was stimulated by intraliposomal ATP and nearly independent of the pH. The K(m) for carnitine was 36 μM. [(3)H]carnitine transport was inhibited by carnitine analogues and some Cys and NH(2) reagents. This paper represents the first outcome in over-expressing, in active form, the third member of the OCTN sub-family, mOCTN3, in E. coli.     

Ochratoxin A adsorption phenotype: An inheritable yeast trait

This study aimed to evaluate the inheritance of the trait ochratoxin A adsorption in two wine strains of Saccharomyces cerevisiae and their 46 descendants. Each strain was inoculated in triplicate in test tubes containing 10 ml of must obtained from the Calabrian Zibibbo white grape variety, artificially contaminated with ochratoxin A to reach a total content of 4.10 ng/ml. The microvinification trials were performed at 25°C. After 30 days, ochratoxin A values ranged from 0.74 to 3.18 ng/ml, from 0.01 to 2.69 ng/ml, and from 0.60 to 2.95 ng/ml respectively in wines, in lees after washing, and in the saline solution used to wash the lees. The analysis of OTA in wines was performed to find the residual toxin content after yeast activity, thus obtaining technological evidence of yeast influence on wine detoxification. The analysis of OTA in lees after washing was performed to distinguish the OTA linked to cells. The analysis of OTA in the saline solution used to wash the lees was performed to distinguish the OTA adsorbed on yeast cell walls and removed by washing, thus focusing on the adsorption activity of wine yeast through electrostatic and ionic interactions between parietal mannoproteins and OTA. Ploidy of the two parental strains was controlled by flow cytometry. Results demonstrated that the ochratoxin A adsorption is genetically controlled and is a polygenic inheritable trait of wine yeasts. The majority of the descendants are characterized by a great and significant diversity compared to their parents. Both the parental strains had genome sizes consistent with their being diploid, so validating the observed results. These findings constitute an initial step to demonstrate the mechanisms of inheritance and establish breeding strategies to improve the ochratoxin A adsorption trait in wine yeasts. This will allow a decrease in the ochratoxin A content of contaminated musts during winemaking, by using genetically improved wine yeasts.     

Outdoor H₂ production in a 50-L tubular photobioreactor by means of a sulfur-deprived culture of the microalga Chlamydomonas reinhardtii

In the past decade, H? production using the green microalga Chlamydomonas reinhardtii has been extensively studied under laboratory-scale photobioreactors, while information on outdoor cultures is still lacking. In this paper, the results of experiments conducted with sulfur-deprived cultures of C. reinhardtii carried out in a 50-L horizontal tubular photobioreactor are presented. Hydrogen production experiments were carried out under both artificial and direct solar light. In both cases, the H? output attained was 18-20% of what obtained in the laboratory. However, no significant changes in the H? production were observed when cells grown outdoors were tested under laboratory conditions. Chlorophyll fluorescence measurements showed that outdoor cultures were subjected to strong photo-inhibition, due to the combination of high solar light intensity and sulfur-deprivation. Indeed, H? production was only achieved outdoors when cultures were previously acclimated to sunlight, a condition that caused a number of physiological changes, namely: (i) a decrease in the chlorophyll content per unit of dry weight; (ii) an increase in the photosynthesis and respiration rates, and (iii) a higher induction of the xanthophyll cycle pigments as compared to non-acclimated cultures. It was concluded that the reduced H? output achieved in the 50-L photobioreactor was due to the different illumination pattern to which the cultures were exposed (one-sided vs. two-sided illumination provided in the laboratory), as well as to the great difference in the mixing times (60 min vs. 15.5s achieved in the lab-scale photobioreactor). To the very best of our knowledge this is the first time that H? production with green algae has been achieved by means of solar light.     

The Di Bella Method (DBM) improved survival, objective response and performance status in a retrospective observational clinical study on 23 tumours of the head and neck

In 23 cases of carcinoma of the head and neck, the combined use of Somatostatin and/or its analogue Octreotide, prolactin inhibitors, Melatonin, Retinoids, Vitamin E, Vitamin D3, Vitamin C, Calcium, chondroitin-sulphate, and minimal oral doses of cyclophosphamide (50-100 mg/day) led to a decided increase in survival with respect to the median values reported in the literature for the same tumours and stages, together with an evident improvement in the quality of life, partial or complete objective responses and, in some cases, complete and stable cure with functional recovery. The rationale and the mechanisms of molecular biology of the treatment are discussed, showing that the treatment has a differentiating, apoptotic, antiproliferative, antiangiogenic and antimetastatic effect, and, unlike chemo- and/or radiotherapy, preserves and enhances the trophism and functionality of organs, tissues and immunitary and antitumoral homeostasis. This result, achieved without toxicity, demonstrates the efficacy of this biological multitherapy (Prof. Luigi Di Bella's method or DBM) and is in agreement with the positive results already published on the use of the DBM in various neoplastic diseases. We believe it is of use to report these cases to invite greater interest in the possibilities opened up by this biological multitherapy.     

Oligomers of the ATPase EHD2 confine caveolae to the plasma membrane through association with actin

Caveolae are specialized domains present in the plasma membrane (PM) of most mammalian cell types. They function in signalling, membrane regulation, and endocytosis. We found that the Eps-15 homology domain-containing protein 2 (EHD2, an ATPase) associated with the static population of PM caveolae. Recruitment to the PM involved ATP binding, interaction with anionic lipids, and oligomerization into large complexes (60-75S) via interaction of the EH domains with intrinsic NPF/KPF motifs. Hydrolysis of ATP was essential for binding of EHD2 complexes to caveolae. EHD2 was found to undergo dynamic exchange at caveolae, a process that depended on a functional ATPase cycle. Depletion of EHD2 by siRNA or expression of a dominant-negative mutant dramatically increased the fraction of mobile caveolar vesicles coming from the PM. Overexpression of EHD2, in turn, caused confinement of cholera toxin B in caveolae. The confining role of EHD2 relied on its capacity to link caveolae to actin filaments. Thus, EHD2 likely plays a key role in adjusting the balance between PM functions of stationary caveolae and the role of caveolae as vesicular carriers.     

On the Evolutionary Developmental Biology of Speciation

The mainstream approaches to the study of speciation and clade diversification have extensively focused on genetic mechanisms and ecological contexts, while much less attention has been paid to the role of development. In this paper we provide materials to support the thesis that taking development into the picture of evolutionary processes can bring important insights on how species multiply and diversify. Evidence that developmentally entangled evolutionary factors are important in speciation comes from different lines of investigation that can be broadly grouped under three headings: evolvability, phenotypic plasticity, and phenology. Evolvability enters the scene through the complexity of the genotype-phenotype map, the developmental link between transmissible genetic information and selectable phenotypes. Phenotypic plasticity can act as a facilitator for speciation, promoting diversification at different stages of the speciation process, as well as generating novel targets and novel trade-offs for evolutionary processes. The formal inclusion of the developmental time axis in speciation models widens the scope for investigating the onset and/or reinforcement of reproductive barriers through a range of situations along an organism??s life cycle. Overall, developmental processes can contribute to speciation and diversification at different stages of the speciation process, at different levels of biological organization and along the organism??s whole life cycle.