Use of the KlADH3 promoter for the quantitative production of the murine PDE5A isoforms in the yeast Kluyveromyces lactis

Background: Phosphodiesterases (PDE) are a superfamily of enzymes that hydrolyse cyclic nucleotides (cAMP/ cGMP), signal molecules in transduction pathways regulating crucial aspects of cell life. PDEs regulate the intensity and duration of the cyclic nucleotides signal modulating the downstream biological efect. Due to this critical role associated with the extensive distribution and multiplicity of isozymes, the 11 mammalian families (PDE1 to PDE11) constitute key therapeutic targets. PDE5, one of these cGMP-specifc hydrolysing families, is the molecular target of several well known drugs used to treat erectile dysfunction and pulmonary hypertension. Kluyveromyces lactis, one of the few yeasts capable of utilizing lactose, is an attractive host alternative to Saccharomyces cerevisiae for heterologous protein production. Here we established K. lactis as a powerful host for the quantitative production of the murine PDE5 isoforms. Results: Using the promoter of the highly expressed KlADH3 gene, multicopy plasmids were engineered to produce the native and recombinant Mus musculus PDE5 in K. lactis. Yeast cells produced large amounts of the purifed A1, A2 and A3 isoforms displaying Km, Vmax and Sildenafl inhibition values similar to those of the native murine enzymes. PDE5 whose yield was nearly 1 mg/g wet weight biomass for all three isozymes (30 mg/L culture), is well tolerated by K. lactis cells without major growth defciencies and interferences with the endogenous cAMP/cGMP signal transduction pathways. Conclusions: To our knowledge, this is the frst time that the entire PDE5 isozymes family containing both regulatory and catalytic domains has been produced at high levels in a heterologous eukaryotic organism. K. lactis has been shown to be a very promising host platform for large scale production of mammalian PDEs for biochemical and structural studies and for the development of new specifc PDE inhibitors for therapeutic applications in many pathologies

Los problemas resueltos en textos universitarios de física

This study presents a description of Physics university solved problems textbooks. The aim is to make a characterisation of authors' treatment of the presentation and resolution of these problems and to investigate its congruity with an investigation model of problem solving. From the analysis of the results it comes out that a presentation of the problematic situation as a closed problem predominates in the sample. Also basic problem-solving processes are not explicit and analysis of results is limited. It is concluded that treatment of solved problems textbooks as examples presents weak coherence with a scientific conception of problem solving

k-dimensional Size Functions for shape description and comparison

This paper advises the use of k-dimensional size functions for comparison and retrieval in the context of multidimensional shapes, where by shape we mean something in two or higher dimensions having a visual appearance. The attractive feature of k-dimensional size functions is that they allow to readily establish a similarity measure between shapes of arbitrary dimension, taking into account different properties expressed by a multivalued real function defined on the shape. This task is achieved through a particular projection of k-dimensional size functions into the 1-dimensional case. Therefore, previous results on the stability for matching purposes become applicable to a wider range of data. We outline the potential of our approach in a series of experiments

The saponification of lauric acid with calcium soaps as an alternative to in-feed antibiotics in post-weaning piglets

In the search for the reduction of antibiotics in farm animals, a concept was developed based on studies with medium-chain fatty acid with 6-12 atoms (MCFA). In particular, they have been shown to exhibit against Gram+ bacteria in piglets at relatively high concentrations. However they can be hardly used as such because of their repellent odour and taste and for their rapid absorption in upper gastrointestinal tract. These problems could be overcome by the generation of monoacylglycerol, but esterification is usually carried out on a silica base, which reduces the concentration of FA, therefore limiting the antibacterial effects. Our hypothesis is that the saponification with calcium salts might positively affect their concentration in the GIT. The aim of the present study was to examine the effects of Laurate calcium soap (C12-Ca soap) on growth performance and health of post-weaning piglets. At weaning, 192 crossbreed Topics piglets were assigned to 3 experimental groups consisting of 16 pens (4 pigs/pen each): CTR (negative control), T1 (basal diet plus Amoxycillin at 400 mg/kg), and T2 (basal diet plus C12-Ca soap at 1 kg/ton). Gain and feed consumption did not differ among groups. Feed efficiency was higher in T1 (0,61) and T2 (0,58) than CTR (0,51) (P<0,01). Mortality was 0 in T1, and reduced in T2 (4,7%) compared to CTR (10,9%). These preliminary results suggest that saponification of MCFA may be a valuable alternative to in-feed antibiotics, used for growth promotion, and even for enhancing health in post-weaning piglets

Approaches for Future Internet architecture design and Quality of Experience (QoE) Control

Researching a Future Internet capable of overcoming the current Internet limitations is a strategic investment. In this respect, this paper presents some concepts that can contribute to provide some guidelines to overcome the above-mentioned limitations. In the authors' vision, a key Future Internet target is to allow applications to transparently, efficiently and flexibly exploit the available network resources with the aim to match the users' expectations. Such expectations could be expressed in terms of a properly defined Quality of Experience (QoE). In this respect, this paper provides some approaches for coping with the QoE provision problem

Un estudio sobre el rozamiento en libros de física de nivel universitario

This work analyses the treatment that is given to friction phenomena in a sample of fifteen physics textbooks for undergraduate level. From this analysis it emerges that friction phenomena is treated fragmentarily. It is associated --almost exclusively-- either with dissipating effects or to delaying forces, it is explained using different models for different scales, systems, materials and surfaces conditions, and clear domain limits for empirical laws are not always established, showing inconsistency in the treatment of the subject. The content analysis technique and linguistic analysis were used in this study. The results are interpreted in the frame of the Rosch prototype theory

Be as Careful of the Company You Keep as of the Books You Read: Peer Effects in Education and on the Labor Market

In this paper we investigate whether peers' behavior influences the choice of college major, thus contributing to the mismatch of skills in the labor market. Using a newly constructed dataset, we are able to identify the endogenous effect of peers on such decisions through a novel identification strategy that solves the common econometric problems of studies of social interactions. Results show that, indeed, one is more likely to choose a major when many of her peers make the same choice. We also provide evidence on skills mismatch in terms of entry wages and occupation. We find that peers can divert students from majors in which they have a relative ability advantage, with adverse consequences on academic performance, entry wages and job satisfaction.

Approximating the 2-dimensional matching distance

Some new approximation results about the 2-dimensional matching distance are presented, leading to the formulation of an algorithm for its computation (up to an arbitrary input error)

Exceptional and Durable Responses to TDM-1 After Trastuzumab Failure for Breast Cancer Skin Metastases: Potential Implications of an Immunological Sanctuary

Breast Cancer (BC) skin metastases represent a challenging clinical scenario. Although they usually arise when other distant metastases are already present, they may also represent a form of locoregional recurrence (LRR). Systemic therapy in this setting may have a role both in case a radical locoregional approach is unfeasible in order to achieve disease control, and as adjuvant strategy after radical removal of cutaneous lesions, in order to prevent or delay subsequent disease spread. Systemic therapy for HER2+ metastatic BC (MBC) currently relies on anti-HER2 targeted agents. In this context TDM1 is an option in trastuzumab-resistant patients.Here we present 2 cases of isolated skin metastases in patients with HER2+ BC progressing during or early after trastuzumab-based therapy, showing impressive responses to TDM1. We hypothesize that the unique properties of skin immune microenvironment may explain the failure of trastuzumab, which exerts its action also through immunological mechanisms, and the subsequent outlier responses to TDM1, that relies on a partially different mechanism of action

Approximating the 2-dimensional matching distance

Some new approximation results about the 2-dimensional matching distance are presented, leading to the formulation of an algorithm for its computation (up to an arbitrary input error)