Human recombinant domain antibodies against multiple sclerosis antigenic peptide CSF114(Glc)

Multiple sclerosis (MS) is a chronic auto‐immune disease characterized by a damage to the myelin component of the central nervous system. Self‐antigens created by aberrant glycosylation have been described to be a key component in the formation of auto‐antibodies. CSF114(Glc) is a synthetic glucopeptide detecting in vitro MS‐specific auto‐antibodies, and it is actively used in diagnostics and research to monitor and quantify MS‐associated Ig levels. We reasoned that antibodies raised against this probe could have been relevant for MS. We therefore screened a human Domain Antibody library against CSF114(Glc) using magnetic separation as a panning method. We obtained and described several clones, and the one with the highest signals was produced as a 6×His‐tagged protein to properly study the binding properties as a soluble antibody. By surface plasmon resonance measurements, we evidenced that our clone recognized CSF114(Glc) with high affinity and specific for the glucosylated peptide. Kinetic parameters of peptide–clone interaction were calculated obtaining a value of K_D in the nanomolar range. Harboring a human framework, this antibody should be very well tolerated by human immune system and may represent a valuable tool for MS diagnosis and therapy, paving the way to new research strategies. Copyright © 2014 John Wiley & Sons, Ltd.     

Efficient optimization of pyrazolo[3,4-d]pyrimidines derivatives as c-Src kinase inhibitors in neuroblastoma treatment

The proto-oncogene c-Src is a non-receptor tyrosine kinase which is involved in the regulation of many cellular processes, such as differentiation, adhesion and survival. c-Src hyperactivation has been detected in many tumors, including neuroblastoma (NB), one of the major causes of death from neoplasia in infancy. We already reported a large family of pyrazolo[3,4-d]pyrimidines active as c-Src inhibitors. Interestingly, some of these derivatives resulted also active on SH-SY5Y NB cell line. Herein, starting from our previous Free Energy Perturbation/Monte Carlo calculations, we report an optimization study which led to the identification of a new series of derivatives endowed with nanomolar K_i values against c-Src, interesting antiproliferative activity on SH-SY5Y cells and a suitable ADME profile.     

Mutations of cytochrome b6 in Chlamydomonas reinhardtii disclose the functional significance for a proline to leucine conversion by petB editing in maize and tobacco

We have introduced a proline codon in place of a leucine codon at position 204 of the petB gene of Chlamydomonas reinhardtii. This gene modification mimics the presence of proline codons at the same position in the petB genes of maize and tobacco, which are subsequently edited to leucine codons at the RNA level. Following transformation, we observed no editing at this position in C. reinhardtii, independent of the type of proline codon we have used: the CCA codon, edited in maize, or a CCT codon. Strains carrying the introduced mutation were non phototrophic and displayed a block in photosynthetic electron transfer, consistent with a lack of cytochrome b6f activity. Thus the presence of a proline residue at position 204 in cytochrome b6 is detrimental to photosynthesis. We show that the mutant phenotype arose from a defective assembly of cytochrome b6f complexes and not from altered electron transfer properties in the assembled protein complex. Biochemical comparison of the proline-containing transformants with a cytochrome b6 mutant deficient in heme-attachment indicates that their primary defect is at the level of assembly of apocytochrome b6 with the bh heme, thereby preventing assembly of the whole cytochrome b6f complex.     

The effect of 50 years of landscape change on species richness and community composition

We analysed a 50-year dataset of avian species observations to determine how richness and community composition varied over a period of landscape-scale environmental change. Our study area, northern lower Michigan, has experienced substantial land-use and land-cover change over time. Like much of the northern Midwest, it has shifted from a largely unpopulated, post-logging shrubland to a moderately populated closed-canopy forest. Such changes are generally expected to influence overall richness and community composition. We found that regional richness per year remained virtually unchanged over the study period. Year-to-year variation in species number was surprisingly low. Richness totals included vastly different species groups as the composition of the regional bird community changed substantially over time. Changes in the types of species present appear to reflect deterministic changes in habitat. The number of grassland and open-habitat species decreased, for example, while species associated with older forests and urban habitats increased. Our results suggest that habitat changes at the landscape scale do not necessarily lead to changes in the number of species a region can support. Such changes, however, do appear to influence the types of species that will occupy a region, and can lead to substantial changes in community composition.     

Staphylococcus aureus Protoplasting Induced by d-Cycloserine

Addition of sublethal doses of d-cycloserine to growing cells of Staphylococcus aureus induces the rupture of the cell wall along an equatorial ring, thus allowing the liberation of protoplasts.