Self-assembly in amphiphilic spherical brushes.

The structure of amphiphilic spherical brushes, consisting of the nano-SiO2 core, the hyperbranched polyamidoamine subshell, and a grafted layer of long hydrophobically modified polyacrylamide (HMPAM) chains, in aqueous solution was analyzed and described in the framework of the original mean-field approach. The scaling estimations of the hydrodynamic radius of such polymer brushes as a function of the number of grafted macromolecules allow concluding that the HMPAM shells are in a globular state and that the region of the stretched chains adjacent to the grafting surface is a minor part of the grafted macromolecules and does not have a significant impact on the self-assembly of the HMPAM shell caused by the complex hydrophobic-hydrophilic composition of their monomer units. In mean-field theory, the amphiphilic nature of HMPAM was taken into account by attaching the hydrophobic side group H to some fraction of monomer units of the hydrophilic P backbone. The strong attraction of H groups causes the aggregation of macromolecules, whereas the affinity of hydrophilic P groups to solvent forces the aggregates to increase their surface. Due to such effective surface activity, in poor solvent, the grafted amphiphilic macromolecules could form a spherical compacted structure around the nanoparticle or self-assemble into a "hedgehog" structure with several "spines" having hydrophobic core and hydrophilic shell. State diagrams, obtained theoretically, reveal that the "hedgehog" structure is preferable for a wide range of energetic parameters.

Role of the conserved cysteine residues of the 11.5 kDa subunit in complex I catalytic properties.

Mitochondrial complex I exists as a mixture of two inter-convertible forms: active (A) and de-activated (D), the latter being sensitive to SH-modifying compounds. To investigate if the conserved cysteine-rich 11.5 kDa subunit of Neurospora crassa complex I is involved in this process, we subjected the corresponding genomic DNA to site-directed mutagenesis. The four cysteine residues of the subunit were separately substituted with serine residues and the resulting proteins were independently expressed in a null-mutant strain. All of the obtained mutant strains were able to assemble a complex I with similar kinetic properties to those observed in the wild-type enzyme, indicating that none of the cysteine residues of the 11.5 kDa protein is individually relevant for the A/D transition process. Diminished amounts of assembled complex I seem to be the major effect of these specific mutations. The cysteine residues are likely important to the acquisition and stabilization of the correct 11.5 kDa protein conformation and this is reflected in the assembly/stability of complex I.

New findings of Neurospora in Europe and comparisons of diversity in temperate climates on continental scales.

The life cycles of the conidiating species of Neurospora are adapted to respond to fire, which is reflected in their natural history. Neurospora is found commonly on burned vegetation from the tropic and subtropical regions around the world and through the temperate regions of western North America. In temperate Europe it was unknown whether Neurospora would be as common as it is in North America because it has been reported only occasionally. In 2003 and 2004 a multinational effort surveyed wildfire sites in southern Europe. Neurospora was found commonly from southern Portugal and Spain (37 degrees N) to Switzerland (46 degrees N). Species collected included N. crassa, N. discreta, N. sitophila and N. tetrasperma. The species distribution and spatial dynamics of Neurospora populations showed both similarities and differences when compared between temperate Europe and western North America, both regions of similar latitude, climate and vegetation. For example the predominant species in western North America, N. discreta phylogenetic species 4B, is common but not predominant in Europe, whereas species rare in western North America, N. crassa NcB and N. sitophila, are much more common in Europe. The meiotic drive element Spore killer was also common in European populations of N. sitophila and at a higher proportion than anywhere else in the world. The methods by which organisms spread and adapt to new environments are fundamental ecosystem properties, yet they are little understood. The differences in regional diversity, reported here, can form the basis of testable hypotheses. Questions of phylogeography and adaptations can be addressed specifically by studying Neurospora in nature.

The 29.9 kDa subunit of mitochondrial complex I is involved in the enzyme active/de-active transitions.

Mitochondrial respiratory chain complex I undergoes transitions from active to de-activated forms. We have investigated the phenomenon in sub-mitochondrial particles from Neurospora crassa wild-type and a null-mutant lacking the 29.9 kDa nuclear-coded subunit of complex I. Based on enzymatic activities, genetic crosses and analysis of mitochondrial proteins in sucrose gradients, we found that about one-fifth of complex I with catalytic properties similar to the wild-type enzyme is assembled in the mutant. Mutant complex I still displays active/de-active transitions, indicating that other proteins are involved in the phenomenon. However, the kinetic characteristics of complex I active/de-active transitions in nuo29.9 differ from wild-type. The spontaneous de-activation of the mutant enzyme is much slower, implicating the 29.9 kDa polypeptide in this event. We suggest that the fungal 29.9 kDa protein and its homologues in other organisms may modulate the active/de-active transitions of complex I.

Neurospora strains harboring mitochondrial disease-associated mutations in iron-sulfur subunits of complex I.

We subjected the genes encoding the 19.3-, 21.3c-, and 51-kDa iron-sulfur subunits of respiratory chain complex I from Neurospora crassa to site-directed mutagenesis to mimic mutations in human complex I subunits associated with mitochondrial diseases. The V135M substitution was introduced into the 19.3-kDa cDNA, the P88L and R111H substitutions were separately introduced into the 21.3c-kDa cDNA, and the A353V and T435M alterations were separately introduced into the 51-kDa cDNA. The altered cDNAs were expressed in the corresponding null-mutants under the control of a heterologous promoter. With the exception of the A353V polypeptide, all mutated subunits were able to promote assembly of a functional complex I, rescuing the phenotypes of the respective null-mutants. Complex I from these strains displays spectroscopic and enzymatic properties similar to those observed in the wild-type strain. A decrease in total complex I amounts may be the major impact of the mutations, although expression levels of mutant genes from the heterologous promoter were sometimes lower and may also account for complex I levels. We discuss these findings in relation to the involvement of complex I deficiencies in mitochondrial disease.

Composition of complex I from Neurospora crassa and disruption of two "accessory" subunits.

Respiratory chain complex I of the fungus Neurospora crassa contains at least 39 polypeptide subunits, of which 35 are conserved in mammals. The 11.5 kDa and 14 kDa proteins, homologues of bovine IP15 and B16.6, respectively, are conserved among eukaryotes and belong to the membrane domain of the fungal enzyme. The corresponding genes were separately inactivated by repeat-induced point-mutations, and null-mutant strains of the fungus were isolated. The lack of either subunit leads to the accumulation of distinct intermediates of the membrane arm of complex I. In addition, the peripheral arm of the enzyme seems to be formed in mutant nuo14 but, interestingly, not in mutant nuo11.5. These results and the analysis of enzymatic activities of mutant mitochondria indicate that both polypeptides are required for complex I assembly and function.

Parameters describing motion in carotid artery plaques from ultrasound examination: A reproducibility study.

We have previously developed a method for quantifying motion in carotid artery plaques from sequences of ultrasound (US) radiofrequency images. Here, we examine the intraoperator reproducibility of the results. Five independent recordings were made on each of six symptomatic and six asymptomatic patients, and processed off-line into 29 motion parameters, representing motion amplitude, stretch/compression and shear motion. For the statistical analysis, we used a linear mixed model and investigated the parameters for contributions from individual patients, contributions from recordings on each patient and contributions from heart cycles within each recording. The model was valid for seven parameters calculated over the entire heart cycle (four calculated over the systole only), which all showed good reproducibility (intraclass coefficient for variance over all patients rho(alpha) >/= 0.4). Averaging three recordings of two heart cycles each gives acceptable accuracy (normalised variance of patient means lambda < 0.3). This acquisition scheme is reasonable in a clinical situation.