Living-floors and structures from the Lower Paleolithic to the Bronze Age in Italy

New researches have been performed on the analysis of some Italian dwelling structures dating from the Lower Paleolithic to Bronze Age. Different methods have been applied to each study according to the extensions of the areas explored. The following sites have been analyzed: Isernia La Pineta (Molise), Visogliano (Trieste) - Lower Paleolithic; Grotta del Cavallo (Lecce), Grotta Grande and Riparo del Molare (Salerno) - Middle Paleolithic; Grotta di Fumane (Verona), Riparo Tagliente (Verona), Grotta Continenza (Fucino L'Aquila), San Bartolomeo (Maiella Mountain, Abruzzo) - Upper Paleolithic; Mondeval de Sora (Belluno), Alpe Veglia (Verbania) and Grotta Edera (Aurisina, Trieste) -Mesolithic; Cala Giovanna Piano (Pianosa Island, Livorno), Contraguda (Perfugas, Sassari), Colle Santo Stefano (Fucino, L'Aquila), Catignano (Pescara), Settefonti (L'Aquila) - Neolithic; Castellaro Lagusello (Monzambano, Mantua) - Bronze Age.     

A single in vivo-selected point mutation in the active center of Toxoplasma gondii ferredoxin-NADP+ reductase leads to an inactive enzyme with greatly enhanced affinity for ferredoxin

Electron transfer between plant-type [2Fe-2S] ferredoxin (Fd) and ferredoxin-NADP+ reductase (FNR) depends on the physical interaction between both proteins. We have applied a random mutagenesis approach with subsequent in vivo selection using the yeast two-hybrid system to obtain mutants of Toxoplasma gondii FNR with higher affinity for Fd. One mutant showed a 10-fold enhanced binding using affinity chromatography on immobilized Fd. A single serine-to-arginine exchange in the active site was responsible for its increased affinity. The mutant reductase was also enzymatically inactive. Homology modeling of the mutant FNR-Fd complex predicts substantial alterations of protein-FAD interactions in the active site of the enzyme with subsequent structural changes. Collectively, for the first time a point mutation in this important class of enzymes is described which leads to greatly enhanced affinity for its protein ligand.     

ICBS: a database of interactions between protein chains mediated by beta-sheet formation

MOTIVATION: Interchain beta-sheet (ICBS) interactions occur widely in protein quaternary structures, interactions between proteins and protein aggregation. These interactions play a central role in many biological processes and in diseases ranging from AIDS and cancer to anthrax and Alzheimer's. RESULTS: We have created a comprehensive database of ICBS interactions that is updated on a weekly basis and allows entries to be sorted and searched by relevance and other criteria through a simple Web interface. We derive a simple ICBS index to quantify the relative contributions of the beta-ladders in the overall interchain interaction and compute first- and second-order statistics regarding amino acid composition and pairing at different relative positions in the beta-strands. Analysis of the database reveals a 15.8% prevalence of significant ICBS interactions, the majority of which involve the formation of antiparallel beta-sheets and many of which involve the formation of dimers and oligomers. The frequencies of amino acids in ICBS interfaces are similar to those in intrachain beta-sheet interfaces. A full range of non-covalent interactions between side chains complement the hydrogen-bonding interactions between the main chains. Polar amino acids pair preferentially with polar amino acids and non-polar amino acids pair preferentially with non-polar amino acids among antiparallel (i, j) pairs. We anticipate that the statistics and insights gained from the database will guide the development of agents that control interchain beta-sheet interactions and that the database will help identify new protein interactions and targets for these agents. AVAILABILITY: The database is available at: http://www.igb.uci.edu/servers/icbs/     

Role of the phosphorolysis of deoxyadenosine in the cytotoxic effect of the combination of deoxyadenosine and deoxycoformycin on a human colon carcinoma cell line (LoVo)

In LoVo cells, phosphorolytic activity acting on deoxyadenosine plays a major role in the resistance to the cytotoxic effect of the combination of deoxynucleoside with deoxycoformycin. In fact, the observed dependence of toxicity on cell density appears to be related to the metabolic conversion of deoxyadenosine into adenine. The phosphorylation of the deoxynucleoside, which represents the first step towards the formation of the cytotoxic agent dATP, proceeds at a significantly lower rate as compared to the phosphorolysis of deoxyadenosine. The analysis of the levels of deoxyadenosine and its derivatives in the incubation media reveals that the rates of disappearance of deoxyadenosine and of formation of adenine increase in concert with the reduction of the effect on cell survival.     

Relationship between <Emphasis Type="Italic">Psoroptes cuniculi</Emphasis> and the Internal Bacterium <Emphasis Type="Italic">Serratia marcescens</Emphasis>

The bacterium Serratia marcescens isolated from surface-sterilised Psoroptes cuniculi was found sensitive to the antibiotic Amikacin. Mites placed in this antibiotic for 48–72 h and then washed by centrifugation were found to be alive and S. marcescens-free. Two experimental infestations were undertaken in order to verify the ability of the S. marcescens-free mites to infect and to give ear skin lesions in healthy rabbits and to evaluate the differential ability of the S. marcescens-free and S. marcescens-infected mites to give ear skin lesions. All rabbits were found to be infested, but only rabbits infested with S. marcescens-free mites presented crusts in their ears, whereas mites and/or eggs were only detected in the ear cerumen of all rabbits infested with S. marcescens-infected mites. S. marcescens was isolated only from P. cuniculi mites taken from these latter rabbits. Results indicate that P. cuniculi mites do not need S. marcescens to live and to be able to infest a healthy rabbit. In addition, S. marcescens was not isolated from eggs and newly born larvae of S. marcescens-infected P. cuniculi, demonstrating that in a population of P. cuniculi this bacterium is not transmitted transovarially.     

Identification and biochemical characterization of the Anopheles gambiae 3-hydroxykynurenine transaminase

Spontaneous oxidation of 3-hydroxykynureine (3-HK), a metabolic intermediate of the tryptophan degradation pathway, elicits a remarkable oxidative stress response in animal tissues. In the yellow fever mosquito Aedes aegypti the excess of this toxic metabolic intermediate is efficiently removed by a specific 3-HK transaminase, which converts 3-HK into the more stable compound xanthurenic acid. In anopheline mosquitoes transmitting malaria, xanthurenic acid plays an important role in Plasmodium gametocyte maturation and fertility. Using the sequence information provided by the Anopheles gambiae genome and available ESTs, we adopted a PCR-based approach to isolate a 3-HK transaminase coding sequence from the main human malaria vector A. gambiae. Tissue and developmental expression analysis revealed an almost ubiquitary profile, which is in agreement with the physiological role of the enzyme in mosquito development and 3-HK detoxification. A high yield procedure for the expression and purification of a fully active recombinant version of the protein has been developed. Recombinant A. gambiae 3-HK transaminase is a dimeric pyridoxal 5'-phosphate dependent enzyme, showing an optimum pH of 7.8 and a comparable catalytic efficiency for both 3-HK and its immediate catabolic precursor kynurenine. This study may be useful for the identification of 3-HK transaminase inhibitors of potential interest as malaria transmission-blocking drugs or effective insecticides.     

Flavonoids from Lonchocarpus latifolius roots

From the petrol extract of Lonchocarpus latifolius roots, 10 flavonoids were isolated. These included: 3,5-dimethoxy-2',2'-dimethylpyrano-(5',6':8,7)-flavone, 3-methoxy-(2',3':7,8)-furanoflavanone, 3',4'-methylenedioxy-(2',3':7,8)-furanoflavanone, and (2,3-trans-3,4-trans)-3,4-dimethoxy-(2',3':7,8)-furanoflavan, as well as the previously known karanjachromene, karanjin, lanceolatin B, pongachromene, pongaglabrone and ponganpin. Only nine flavonoids could be quantified through HPLC analysis.     

Identification and characterization of heart-specific splicing of human neurexin 3 mRNA (NRXN3)

Three neurexin (NRXN) genes are known in humans, each transcribed from two promoters and extensively spliced at five canonical positions, thus generating thousands of isoforms. For NRXN3, only neuronal expression was reported so far. We reported here on the expression of NRXN3 in additional tissues (lung, pancreas, heart, placenta, liver, and kidney) and on the identification and characterization of heart-specific splicing variants of NRXN3. Cardiac isoforms of NRXN3 probably participate in a complex involving dystroglycan and proteins of extracellular matrix, involved in intercellular connections.     

Improving the prediction of protein secondary structure in three and eight classes using recurrent neural networks and profiles

Secondary structure predictions are increasingly becoming the workhorse for several methods aiming at predicting protein structure and function. Here we use ensembles of bidirectional recurrent neural network architectures, PSI-BLAST-derived profiles, and a large nonredundant training set to derive two new predictors: (a) the second version of the SSpro program for secondary structure classification into three categories and (b) the first version of the SSpro8 program for secondary structure classification into the eight classes produced by the DSSP program. We describe the results of three different test sets on which SSpro achieved a sustained performance of about 78% correct prediction. We report confusion matrices, compare PSI-BLAST to BLAST-derived profiles, and assess the corresponding performance improvements. SSpro and SSpro8 are implemented as web servers, available together with other structural feature predictors at: http://promoter.ics.uci.edu/BRNN-PRED/.