Balancing crosstalk between 20-hydroxyecdysone-induced autophagy and caspase activity in the fat body during Drosophila larval-prepupal transition

In the fruitfly, Drosophila melanogaster, autophagy and caspase activity function in parallel in the salivary gland during metamorphosis and in a common regulatory hierarchy during oogenesis. Both autophagy and caspase activity progressively increase in the remodeling fat body, and they are induced by a pulse of the molting hormone (20-hydroxyecdysone, 20E) during the larval-prepupal transition. Inhibition of autophagy and/or caspase activity in the remodeling fat body results in 25–40% pupal lethality, depending on the genotypes. Interestingly, a balancing crosstalk occurs between autophagy and caspase activity in this tissue: the inhibition of autophagy induces caspase activity and the inhibition of caspases induces autophagy. The Drosophila remodeling fat body provides an in vivo model for understanding the molecular mechanism of the balancing crosstalk between autophagy and caspase activity, which oppose with each other and are induced by the common stimulus 20E, and blockage of either path reinforces the other path.     

Extracellular complementation of a developmental mutation implicates a small sporulation protein in aerial mycelium formation by S. coelicolor

The filamentous bacterium S. coelicolor differentiates by forming aerial hyphae, which protrude into the air and metamorphose into chains of spores. Aerial hyphae formation is associated with the production of a small, abundant protein, SapB, which is present in a zone around colonies of differentiating bacteria. Production of SapB is impaired in bld mutants, which are blocked in aerial hyphae formation, but not in whi mutants in which spore formation is prevented. We report that aerial hyphae formation by a newly identified bld mutant is restored by juxtaposition of the mutant near colonies of SapB-producing bacteria or by the application of the purified protein near mutant colonies. These observations implicate SapB in aerial mycelium formation and suggest that SapB is a morphogenetic protein that enables hyphae on the surface of colonies to grow into the air.     

Studies on the catalytic mechanism of H2-forming methylenetetrahydromethanopterin dehydrogenase: para-ortho H2 conversion rates in H2O and D2O

H₂–forming N ⁵,N ¹⁰ –methylenetetrahydromethanopterin dehydrogenase is a novel type of hydrogenase that contains neither nickel nor iron-sulfur clusters. Evidence has been presented that the reaction mechanism catalyzed by the enzyme is very similar to that of the formation of carbocations and H₂ from alkanes under superacidic conditions. We present here further results in support of this mechanism. It was found that the purified enzyme per se did not catalyze the conversion of para H₂ to ortho H₂, a reaction catalyzed by all other hydrogenases known to date. However, it catalyzed the conversion in the presence of the substrate N ⁵,N ¹⁰ –methenyltetrahydromethanopterin (CH≡H₄MPT⁺), indicating that for heterolytic cleavage of H₂ the enzyme-CH≡H₄MPT⁺ complex is required. In D₂O, the formation of HD and D₂ from H₂ rather than a para–ortho H₂ conversion was observed, indicating that after heterolytic cleavage of H₂ the dissociation of the proton from the enzyme-substrate complex is fast relative to the re-formation of free H₂.     

New biotech applications from evolved D-amino acid oxidases

D-Amino acid oxidase (DAAO) is a well-known flavoenzyme that catalyzes the oxygen-dependent oxidative deamination of amino acid D-isomers with absolute stereospecificity, which results in α-keto acids, ammonia and hydrogen peroxide. Recently, the extraordinary functional plasticity of DAAO has become evident; in turn, boosting research on this flavoprotein. Protein engineering has allowed for a redesign of DAAO substrate specificity, oxygen affinity, cofactor binding, stability, and oligomeric state. We review recent developments in utilizing DAAO, including as a biocatalyst for resolving racemic amino acid mixtures, as a tool for biosensing, and as a new mechanism of herbicide resistance. Perspectives for future biotechnological applications of this oxidative biocatalyst are also outlined.     

Investigating the role of active site residues of Rhodotorula gracilis D-amino acid oxidase on its substrate specificity

D-amino acid oxidase (DAAO) is a flavoprotein that catalyzes stereospecifically the oxidative deamination of D-amino acids. The wild-type DAAO is mainly active on neutral D-amino acids, while basic D-amino acids are poor substrates and the acidic ones are virtually not oxidized. To present a comprehensive picture of how the active site residues can modulate the substrate specificity a number of mutants at position M213, Y223, Y238, R285, S335, and Q339 were prepared in the enzyme from the yeast Rhodotorula gracilis. All DAAO mutants have spectral properties similar to those of the wild-type enzyme and are catalytically active, thus excluding an essential role in catalysis; a lower activity on neutral and basic amino acids was observed. Interestingly, an increase in activity and (k(cat)/K(m))(app) ratio on D-aspartate was observed for all the mutants containing an additional charged residue in the active site. The active site of yeast DAAO appears to be a highly evolved scaffold built up through evolution to optimize the oxidative deamination of neutral D-amino acids without limiting its substrate specificity. It is noteworthy, that introduction of a sole, additional, positively charged residue in the active site is sufficient to optimize the reactivity on acidic D-amino acids, giving rise to kinetic properties similar to those of D-aspartate oxidase.     

Italian Dermestidae: notes on some species and?an?updated checklist (Coleoptera)

An up-to-date checklist of the Italian Dermestidae is provided. The presence of 95 species in Italy is confirmed, while further 5 species (Dermestes (Dermestes) vorax Motschulsky, 1860, Thorictuspilosus Peyron, 1857, T. wasmanni Reitter, 1895, Attagenus (Attagenus) simonis Reitter, 1881 and Globicornis (G.) breviclavis (Reitter, 1878)) and 1 subspecies (A. (A.) tigrinus pulcher Faldermann, 1835) are excluded from the Italian fauna.Attagenus (Attagenus) calabricus Reitter, 1881 and A. (A.) lobatus Rosenhauer, 1856 are for the first time recorded from Abruzzi and Tuscany respectively; A. (A.) silvaticus Zhantiev, 1976 is recorded for the first time from mainland Italy (Apulia); Anthrenus (Anthrenus) angustefasciatus Ganglbauer, 1904 is new to northern Italy (Friuli-Venezia Giulia), central Italy (Tuscany), Apulia and Basilicata; A. (A.) munroi Hinton, 1943 is new to central Italy (Elba Island); A. (A.) delicatus Kiesenwetter, 1851 is for the first time recorded from Apulia; Globicornis (Globicornis) fasciata (Fairmaire & Brisout de Barneville, 1859) is new to southern Italy (Basilicata); G. (Hadrotoma) sulcata (C.N.F. Brisout de Barneville, 1866) is for the first time recorded from central Italy (Abruzzi), Campania and Sicily, whileTrogoderma inclusum LeConte, 1854 is new to Apulia.Seven species (Dermestes (Dermestes) peruvianus Laporte de Castelnau, 1840, D. (Dermestinus) carnivorus Fabricius, 1775, D. (Dermestinus) hankae Háva, 1999, D. (Dermestinus) intermedius intermedius Kalík, 1951, D. (Dermestinus) szekessyi Kalík, 1950, Anthrenus (Anthrenops) coloratus Reitter, 1881 and Trogodermaangustum (Solier, 1849)) recently recorded from Italy (without further details) are discussed.The lectotype and a paralectotype are designated forAttagenus (A.) calabricus Reitter, 1881 from Calabria.Attagenus pellio (Linnaeus, 1758) var. pilosissimus Roubal, 1932 is removed from synonymy with A. (A.) pellio and recognized as a valid species (stat. prom.); it is known from Lombardy, Apulia and Calabria.     

The cold-active lipase of Pseudomonas fragi. Heterologous expression, biochemical characterization and molecular modeling.

A recombinant lipase cloned from Pseudomonas fragi strain IFO 3458 (PFL) was found to retain significant activity at low temperature. In an attempt to elucidate the structural basis of this behaviour, a model of its three-dimensional structure was built by homology and compared with homologous mesophilic lipases, i.e. the Pseudomonas aeruginosa lipase (45% sequence identity) and Burkholderia cepacia lipase (38%). In this model, features common to all known lipases have been identified, such as the catalytic triad (S83, D238 and H260) and the oxyanion hole (L17, Q84). Structural modifications recurrent in cold-adaptation, i.e. a large amount of charged residues exposed at the protein surface, have been detected. Noteworthy is the lack of a disulphide bridge conserved in homologous Pseudomonas lipases that may contribute to increased conformational flexibility of the cold-active enzyme.     

Inference in dynamic systems using B‐splines and quasilinearized ODE penalties

Nonlinear (systems of) ordinary differential equations (ODEs) are common tools in the analysis of complex one‐dimensional dynamic systems. We propose a smoothing approach regularized by a quasilinearized ODE‐based penalty. Within the quasilinearized spline‐based framework, the estimation reduces to a conditionally linear problem for the optimization of the spline coefficients. Furthermore, standard ODE compliance parameter(s) selection criteria are applicable. We evaluate the performances of the proposed strategy through simulated and real data examples. Simulation studies suggest that the proposed procedure ensures more accurate estimates than standard nonlinear least squares approaches when the state (initial and/or boundary) conditions are not known.     

The role of miRNA-221 and miRNA-126 in patients with benign metastasizing leiomyoma of the lung: an overview with new interesting scenarios

Molecular Biology Reports - Benign metastasizing leiomyoma (BML) is a rare disease characterized by extrauterine benign leiomyomatosis in patients with a previous or concomitant history of uterine...