Determination of D-amino acids using a D-amino acid oxidase biosensor with spectrophotometric and potentiometric detection

An amperometric and a colorimetric biosensor to detect and quantify D-amino acids selectively has been devised using D-amino acid oxidase from Rhodotorula gracilis. The sensor is characterised by a proportional response between 0.2-3 mM and 0.1-1 mM D-alanine for the amperometric (at a working potential of 1400 mV vs Ag/AgCl) and colorimetric system, respectively.     

L'ipotesi dell'effetto del campo geomagnetico e la variabilità tra le ripetizioni in prove di germinazione ed accrescimento in Triticum

Abstract

The geomagnetic field effect hypothesis and the variability among replications in germination and growth tests in Triticum. – During previous eperiments on germination and growth of Triticum, we have often found many cases of heteroscedasticity and of statistically significant differences among contemporary replications (i.e. germinators). In such experiments all the caryopses were oriented in the same way inside the germinators, whilst the germinators were randomly set inside the thermostats: therefore the seeds of different germinators might result differently oriented. Some researchers report that seeds differently oriented according to the lines of force of the geomagnetic field, may respond with different germination, growth rates and growth directions of shoots and roots (magnetotropism). The present investigation was designed to ascertain if these effects might be responsible of the variability among the contemporary replications. The different caryopses orientation within laboratory geomagnetic field, does not give evidence of any effect on germination, growth rate and direction. The caryopses show no germination sensibility neither related to different age, light and temperature conditions, neither to different cultivars, neither to varying orientation from the geomagnetic North every 15° and to varying time of ? preorientation ? in dry conditions. Differences between the variability of randomly and North oriented subsamples are not observed. No statistically significant differences are observed among shoots and roots lenghts of seedlings from differently oriented caryopses neither in different environmental conditions, nor as function of the orientation (24 angles of 15°), nor after different preorientations, nor after repeating the test on 608 subsamples (12.000 seedlings). Therefore the variability among contemporary replications is not imputable to the geomagnetic field. The variability and the distributions structure analysis suggest that both the heteroscedasticity and the significant differences among the replications may be due to an interaction among the caryopses inside the germinators through the substrate. This is probably related to the cultivar characteristics. We emphasize that an accurate evaluation of the variability is particularly necessary in germination and growth tests.     

Xylanase Activity of Phanerochaete chrysosporium

Xylan-degrading enzymes were induced when Phanerochaete chrysosporium was grown at 30°C in shake flask media containing xylan, Avicel PH 102, or ground corn stalks. The highest xylanase activity was produced in the corn stalk medium, while the xylan-based fermentation resulted in the lowest induction. Analytical and preparative isoelectric focusing were used to characterize xylanase multienzyme components. Preparative focusing was performed only with the cultures grown on Avicel and corn stalk. Of over 30 protein bands separated by analytical focusing from the Avicel and corn stalk media, three main groups (I, II, and III) of about five isoenzymes each showed xylanase activity when a zymogram technique with a xylan overlay was used. Enzyme assays revealed the presence of 1,4-β-endoxylanase and arabinofuranosidase activities in all three isoenzyme groups separated by preparative isoelectric focusing. β-Xylosidase activity appeared in the first peak and also as an independent peak between peaks II and III. Denatured molecular masses for the three isoenzyme groups were found to be between 18 and 90 kDa, and pI values were in the range of 4.2 to 6.0. β-Xylosidase has an apparent molecular mass of 20, 30, and 90 kDa (peak I) and 18 and 45 kDa (independent peak), indicating a trimer and dimer structure, respectively, with pI values of 4.2 and 5.78, respectively. Three more minor xylanase groups were produced on corn stalk medium: a double peak in the acidic range (pI 6.25 to 6.65 and 6.65 to 7.12) and two minor peaks in the alkaline range (pI 8.09 to 8.29 and 9.28 to 9.48, respectively). The profile of xylanases separated by isoelectric focusing (zymogram) of culture filtrate from cells grown on corn stalk media was more complex than that of culture supernatants from cells grown on cellulose. The pH optima of the three major xylanase groups are in the range of pH 4 to 5.5.     

Polymorphisms of the prion protein gene in Italian patients with Creutzfeldt-Jakob disease

Creutzfeldt-Jakob disease (CJD) is a transmissible neurodegenerative disorder characterized by the accumulation of the amyloid protein PrP in the CNS. Two coding polymorphisms of the PrP gene (PRNP) are a methionine (Met) to valine (Val) change at codon 129, and a deletion in the octapeptide coding region. In the United Kingdom, homozygosity at codon 129 appears to be associated with a predisposition to develop CJD. However, in Japan, where allelic frequencies and genotype distribution are significantly different, such an association has not been demonstrated. To determine whether such deletion(s) or codon 129 polymorphisms of PRNP predispose to the development of CJD in Italian patients, 31 sporadic CJD patients with no known PRNP mutations, and 186 unrelated control subjects were studied. Genotypic frequencies at codon 129 in these Italian CJD patients revealed a significant excess of methionine alleles, and a different genotype distribution in comparison with the normal Italian population. Deletions of a 24-bp segment located in the PrP octapeptide coding region were found in two control subjects, but in none of the sporadic CJD patients. These data suggest that Met homozygosity at codon 129 may contribute, with other enviromental or endogenous factors, to CJD development.     

Mining cancer biology through bioinformatic analysis of proteomic data

ABSTRACT

Introduction: Discovery proteomics for cancer research generates complex datasets of diagnostic, prognostic, and therapeutic significance in human cancer. With the advent of high-resolution mass spectrometers, able to identify thousands of proteins in complex biological samples, only the application of bioinformatics can lead to the interpretation of data which can be relevant for cancer research.

Areas covered: Here, we give an overview of the current bioinformatic tools used in cancer proteomics. Moreover, we describe their applications in cancer proteomics studies of cell lines, serum, and tissues, highlighting recent results and critically evaluating their outcomes.

Expert opinion: The use of bioinformatic tools is a fundamental step in order to manage the large amount of proteins (from hundreds to thousands) that can be identified and quantified in a cancer biological samples by proteomics. To handle this challenge and obtain useful data for translational medicine, it is important the combined use of different bioinformatic tools. Moreover, a particular attention to the global experimental design, and the integration of multidisciplinary skills are essential for best setting of tool parameters and best interpretation of bioinformatics output.     

Biosynthesis of sulphur amino acids in Saccharomyces cerevisiae

A number of strains of Saccharomyces which produce sulphite by sulphate reduction were examined from an enzymatic and genetic point of view.There are a number of mechanisms that regulate this activity. All of these mechanisms involve the sulphite-reducing activity. In the strains examined, reduced function as a result of mutation in the Sr-locus (affecting H₂S-NADP oxidoreductase EC 1.8.1.2), repression of biosynthesis of the enzyme because of a mutation below the specific locus, and inhibition of the enzyme by endogenous factors were found to be responsible. The production of sulphite can also be connected with a complex state of heterozygosity.It is probably this multiplicity of biochemical and genetic mechanisms that accounts for the frequency with which the production of sulphite is observed in wild strains in nature.This investigation was supported by a research grant of C.N.R. (Consiglio Nazionale delle Ricerche, Roma).