Hydrolysis of fish oil by hyperactivated rhizomucor miehei lipase immobilized by multipoint anion exchange

Rhizomucor miehei lipase (RML) is greatly hyperactivated (around 20‐ to 25‐fold toward small substrates) in the presence of sucrose laurate. Hyperactivation appears to be an intramolecular process because it is very similar for soluble enzymes and covalently immobilized derivatives. The hyperactivated enzyme was immobilized (in the presence of sucrose laurate) on cyanogen bromide‐activated Sepharose (very mild covalent immobilization through the amino terminal residue), on glyoxyl Sepharose (intense multipoint covalent immobilization through the region with the highest amount of Lys residues), and on different anion exchangers (by multipoint anionic exchange through the region with the highest density of negative charges). Covalent immobilization does not promote the fixation of the hyperactivated enzyme, but immobilization on Sepharose Q retains the hyperactivated enzyme even in the absence of a detergent. The hydrolysis of fish oils by these hyperactivated enzyme derivatives was sevenfold faster than by covalently immobilized derivatives and three and a half times faster than by the enzyme hyperactivated on octyl‐Sepharose. The open structure of the hyperactivated lipase is fairly exposed to the medium, and no steric hindrance should interfere with the hydrolysis of large substrates. These new hyperactivated derivatives seem to be more suitable for hydrolysis of oils by RML immobilized inside porous supports. In addition, the hyperactivated derivatives are fairly stable against heat and organic cosolvents. © 2011 American Institute of Chemical Engineers Biotechnol. Prog., 2011     

Novel,achiral aminoheterocycles as selective monoamine reuptake inhibitors

A variety of novel aminoheterocycle scaffolds as selective monoamine reuptake inhibitors have been prepared and one of these scaffolds is achiral. The main elements responsible for hERG channel, CYP2D6 and CYP3A4 inhibition were identified.     

Characterization of selectivity and pharmacophores of type 1 sea anemone toxins by screening seven Na(v) sodium channel isoforms

During their evolution, animals have developed a set of cysteine-rich peptides capable of binding various extracellular sites of voltage-gated sodium channels (VGSC). Sea anemone toxins that target VGSCs delay their inactivation process, but little is known about their selectivities. Here we report the investigation of three native type 1 toxins (CGTX-II, δ-AITX-Bcg1a and δ-AITX-Bcg1b) purified from the venom of Bunodosoma cangicum. Both δ-AITX-Bcg1a and δ-AITX-Bcg1b toxins were fully sequenced. The three peptides were evaluated by patch-clamp technique among Nav1.1-1.7 isoforms expressed in mammalian cell lines, and their preferential targets are Na(v)1.5>1.6>1.1. We also evaluated the role of some supposedly critical residues in the toxins which would interact with the channels, and observed that some substitutions are not critical as expected. In addition, CGTX-II and δ-AITX-Bcg1a evoke different shifts in activation/inactivation Boltzmann curves in Nav1.1 and 1.6. Moreover, our results suggest that the interaction region between toxins and VGSCs is not restricted to the supposed site 3 (S3-S4 linker of domain IV), and this may be a consequence of distinct surface of contact of each peptide vs. targeted channel. Our data suggest that the contact surfaces of each peptide may be related to their surface charges, as CGTX-II is more positive than δ-AITX-Bcg1a and δ-AITX-Bcg1b.     

Immuno-enzymatic evaluation of the recombinant TSSA-II protein of Trypanosoma cruzi in dogs and human sera: a tool for epidemiological studies

The rTSSA-II (recombinant Trypomastigote Small Surface II) antigen was evaluated by ELISA to detect anti-Trypanosoma cruzi antibodies in sera from naturally infected dogs and humans. For this evaluation ELISA-rTSSA-II was standardized and groups were classified according to the results obtained through xenodiagnosis, ELISA and PCR. Sensitivity (Se), Specificity (Sp), Kappa index (KI) and area under curve (AUC) were determined. The Se was determined by using 14 sera from dogs infected with T. cruzi VI (TcVI) whereas Sp was determined by using 95 non-chagasic sera by xenodiagnosis, ELISA-Homogenate and PCR. The performance of ELISA-rTSSA-II in dog sera was high (AUC=0·93 and KI=0·91). The Se was 92·85% (1 false negative) and Sp was 100%. Two sera from dogs infected with TcI and 1 with TcIII were negative. For patients infected with T. cruzi, reactivity was 87·8% (36/41), there was only 1 indeterminate, and Sp was 100%. Fifty-four sera from non-chagasic and 68 sera from patients with cutaneous leishmaniasis did not react with rTSS-II. ELISA-rTSSA-II showed a high performance when studying sera from naturally infected dogs and it also presented 100% Sp. This assay could be an important tool to carry out sero-epidemiological surveys on the prevalence of T. cruzi circulating lineages in the region.     

Hydrolysis of cyclosporin A: Identification of 1,11 seco-cyclosporin A and 4,5 secoisoCyclosporin A by FAB-MS/MS

We have previously reported that treatment of CsA with aqueous HCl gives rise to the formation of a number of water-soluble compounds. Two of these were identified from their FAB-MS/MS spectra as open-chain nona- and decapeptides. We describe here the identification of two other main compounds deriving from the same treatment. Identification was rendered possible from the comparison of their FAB-MS/MS spectra with those of methyl and acetyl derivatives. The two compounds are water-soluble, open-chain undecapeptides corresponding to 1,11 seco-CsA and of 4,5 seco-isoCsA, respectively.     

Inhibition of melanoma development in the Nras(Q61K)::Ink4a−/− mouse model by the small molecule BI‐69A11

To date, there are no effective therapies for tumors bearing NRAS mutations, which are present in 15–20% of human melanomas. Here we extend our earlier studies where we demonstrated that the small molecule BI‐69A11 inhibits the growth of melanoma cell lines. Gene expression analysis revealed the induction of interferon‐ and cell death‐related genes that were associated with responsiveness of melanoma cell lines to BI‐69A11. Strikingly, the administration of BI‐69A11 inhibited melanoma development in genetically modified mice bearing an inducible form of activated Nras and a deletion of the Ink4a gene (Nras^(Q61K)::Ink4a^?/?). Biweekly administration of BI‐69A11 starting at 10 weeks or as late as 24 weeks after the induction of mutant Nras expression inhibited melanoma development (100 and 36%, respectively). BI‐69A11 treatment did not inhibit the development of histiocytic sarcomas, which constitute about 50% of the tumors in this model. BI‐69A11‐resistant Nras^(Q61K)::Ink4a^?/? tumors exhibited increased CD45 expression, reflective of immune cell infiltration and upregulation of gene networks associated with the cytoskeleton, DNA damage response, and small molecule transport. The ability to attenuate the development of NRAS mutant melanomas supports further development of BI‐69A11 for clinical assessment.     

Preface—a new paradigm for life cycle thinking: exploring sustainability in urban development scenarios

The International Journal of Life Cycle Assessment - The main goal of this special issue is to further the understanding of how to integrate life cycle sustainability assessment (LCSA) methods and...     

Water transfer energetics and solid-like packing of globular proteins

In this article we generalize the use of a relationship based on the occurrence of some characteristic temperatures in protein unfolding, which were originally high-lighted for proteins showing the unfolding enthalpy-entropy convergence. On this basis, we show how to dissect the unfolding Gibbs energy of globular proteins in terms of solid-like and liquid-like contributions, untangling the protein energetics by a scheme which does not suffer from excessive intricacy. We also provide an experimental estimate of unfavorable polar contributions to the protein stability, by which it seems possible to evaluate the number of buried residues in individual proteins. A comparison is assessed with the so-called hard sphere model of globular proteins. Results seem to reconcile the view that the protein interior is liquid-like with the observation that protein organization resembles an assembly of closely packed spheres.     

Analytical morphometry of sagittal fronto-facial profiles of 2nd-6th month human foetus

A prominent problem in the morphological study of developing biological structures is shape parametrization with the aim to evaluate transformations in a non subjective way. We carried out an analytical morphometrical study on a series of human embryos, by means of the S.A.M. (Shape Analytical Morphometry) software package. After standardization and normalization of the fronto-facial sagittal profiles, the analytical procedures applied were: 1) Fourier analysis: each profile was considered as an irregular but periodic function obtained by the sum of sinusoids of increasing order. 2) Shape Asymmetry Evaluation: couples of profiles are compared by means of "Janus" procedure; by a parabolic fitting we obtained parameters able to evaluate allometric and isometric differences between the two profiles. The preliminary results of the applied procedures indicated that rate and direction of allometric and isometric growth are not constant during the time.