Cellulose-acetate electrophoresis (CAE) was used to investigate isozyme polymorphisms among different isolates of Fusarium cerealis, F. culmorum, F. graminearum and F. pseudograminearum. After initial testing of 18 enzymes in three buffer systems for activity and resolution of bands, 12 proved to be appropriate for analysis of the full sample set. Comparing the different electrophoretic types (ETs), adenylate kinase (AK), NADP dependent glutamate dehydrogenase (NADP GDH), peptidase B (PEP B), peptidase D (PEP D) and phosphoglucomutase (PGM) proved to be diagnostic for at least one species examined. However, only PEP D was useful alone as a marker to distinguish the four taxa studied providing a rapid and simple CAE based diagnostic protocol.
Biarsenical fluorescein compounds feature unique fluorescence characteristics and special binding mechanism to tetracysteine tags with certain structures and these dyes offer a feasible method for site specific labeling of heterologously expressed proteins. We aimed FlAsH fluorescent labeling of tetracysteine fused hinge region of the ultraspiracle from Drosophila melanogaster (DmUSP-D domain) to facilitate functional studies of this receptor domain. A CCPGCC tetracysteine motif was integrated between His6, Gateway attB1, and Flag tags and attached to the N-terminus of the DmUSP-D. The fusion protein was expressed in Esherichia coli and the FlAsH labeling was performed in bacterial extracts, under conditions which are compatible with receptor function. The dye was bound to the tetracysteine tag with high affinity and complex stability and the labeling proved to be specific for the target fusion protein. Results indicate that FlAsH labeling of the internal CCPGCC motif can be a valuable tool for the functional characterisation of any nuclear receptor domains.
Authors:M. Szécsi, Dóra Ondré, I. Tóth, S. Magony, J. Wölfling, Gy. Schneider, and J. Julesz
The 5α-reductase type 1 isozyme is a key enzyme in the metabolism of the androgen steroid hormones and inhibitors of this enzyme represent a new pharmacological treatment for several androgen dependent diseases. We developed a radiosubstrate
incubation method for the determination of 5α-reductase type 1 activity using rat liver microsomes as an enzyme source. With this method we have studied the inhibiting activity of novel (5′
)-17β-(4,5-dihydrooxazol-5-yl)androst-5-en-3-one compounds containing various derivatized phenyl substituents coupled to the
-heterocyclic moiety. Tests revealed moderate inhibitory actions compared to finasteride, nevertheless, results provide interesting structure-activity relationship data.
Authors:Cs. Dobolyi, F. Sebők, J. Varga, S. Kocsubé, G. Szigeti, N. Baranyi, Á. Szécsi, B. Tóth, M. Varga, B. Kriszt, S. Szoboszlay, C. Krifaton, and J. Kukolya
Climate change affects the occurrence of fungi and their mycotoxins in foods and feeds. A shift has recently been observed in the presence of aflatoxin producer Aspergillus spp. in Europe, with consequent aflatoxin contamination in agricultural commodities including maize in several European countries that have not faced with this problem before, including, e.g. Northern Italy, Serbia, Slovenia, Croatia and Romania. Although aflatoxin contamination of agricultural products including maize is not treated as a serious threat to Hungarian agriculture due to climatic conditions, these observations led us to examine the mycobiota of maize kernels collected from Hungarian maize fields. Using a calmodulin sequence-based approach, A. flavus isolates have been identified in 63.5% of the maize fields examined in 2009 and 2010, and 18.8% of these isolates were found to be able to produce aflatoxins above 5 μg kg−1 on maize kernels as determined by ELISA, HPLC-FL, HPLC-MS analyses and SOS-Chromotest. These data indicate that aflatoxin producing Aspergilli are present in Hungarian agricultural fields, consequently climate change with elevated temperatures could lead to aflatoxin contamination of Hungarian agricultural products, too.