Predicting vegetative inoculum performance to maximize phytase production in solid-state fermentation using response surface methodology

Microbial phytase is used to reduce the environmental loading of phosphorus from animal production facilities. The limiting factors in the use of this enzyme in animal feeds can be overcome by solid-state fermentation (SSF), which is a promising technology for commercial enzyme production with lower production costs. Inoculum quality and the influence of inoculum quality on phytase production are important factors which need in-depth investigation before scaling-up of high-yielding fermentation process. A full factorial experimental design for 240 h with sampling at every 24 h was used to determine the effects of the treatments, inoculum age (plate and liquid culture), media composition and the duration of SSF on the production of fungal biomass and phytase in SSF systems using Aspergillus niger. The optimal treatment combination for maximal phytase production was determined by statistically comparing all treatments at each sampling time. Both 7- and 14-day plate cultures and M1+ medium composition with 72-h-old liquid inoculum treatments resulted in optimal phytase production at 144 h of SSF, which was the shortest duration observed for maximal phytase production. This resulted in maximal phytase production with a mean of 884±121 U/g substrate, while the maximal phytase production observed at 216 h of SSF (mean phytase activity of 1008±121 U/g substrate), with the same treatment combinations, was not statistically significant from that at 144 h of SSF. Phytase production was strongly growth-associated with younger inocula. The significant treatment variables, age of liquid inoculum and the duration of SSF, were used to predict the system response for phytase production using response surface methodology. From the response surface model, the optimal response of the experiment was predicted and the reliability of the prediction was checked with the verification experiment. Journal of Industrial Microbiology & Biotechnology (2001) 26, 161–170. Received 06 June 2000/ Accepted in revised form 14 October 2000     

Overexpression and Purification of an Immunologically Reactive His–BIV Capsid Fusion Protein

The gene of the capsid protein of bovine immunodeficiency virus (BIV) was linked to a sequence encoding for six histidines and expressed as the (His)₆p26 capsid fusion protein. The fusion protein was strongly expressed as both soluble and insoluble forms after induction by isopropylthio-β- -galactoside. Purification was based on interaction of the hexa-histidine polypeptide with metal ions. Expression could represent 11% of the total protein inEscherichia coli,allowing more than 20 mg of highly purified protein to be obtained per liter of bacterial culture. The (His)₆p26 capsid fusion protein purified by immobilized metal affinity chromatography reacted specifically in Western blot with sera from cattle experimentally infected by BIV, as well as with two monoclonal antibodies directed against different epitopes of the Gag protein. The ease of expression, purification, and specificity of this fusion protein should permit a thorough study of prevalence of BIV infection in large-scale serological studies of field samples.     

Computing the selection gradient and evolutionary response of an infinite-dimensional trait

 Following the results developed in a previous paper, an equation describing the evolutionary response to selection is extended from finite- to infinite-dimensional traits. The selection gradient and evolutionary response are then computed for a large class of infinite-dimensional traits of broad biological interest. In this framework, traits are modeled as Gaussian processes, and reproducing kernel Hilbert spaces constitute a primary tool. Received 12 September 1996     

The large bat Helitron DNA transposase forms a compact monomeric assembly that buries and protects its covalently bound 5′-transposon end

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Consistency of growth response to precipitation by Pseudopiptadenia contorta in two Atlantic Forest remnants

Tree growth sensitivity to climate can vary over space and time. This variability generates inconsistency in growth response to climate, which makes it difficult to assess the effects of past climate and global climate change on tree growth. A previous short-term study of Pseudopiptadenia contorta found a consistent growth response to climate in distinct locations, which raises the question, is the growth response of P. contorta to climate consistent over the long-term? We aimed to assess whether there is a common pattern of variation in tree-ring width, build tree-ring width chronologies, and verify the consistency of the climate-growth response of P. contorta in two Atlantic Forest remnants. Wood samples were collected in Reserva Biológica de Poço das Antas (RBPA) and Reserva Biológica de Tinguá (RBT) in the state of Rio de Janeiro, Brazil. Conventional dendrochronology methods were used for cross-dating, to build chronologies and to assess the climate-growth relationship. A common growth pattern was detected for P. contorta, and two tree-ring width chronologies were constructed. A congruent growth response was found for trees of RBPA and RBT to annual and spring precipitation as well as precipitation in the rainy months. Other climate-growth relationships were detected with other precipitation and temperature variables. Considering that P. contorta is a widespread species, occurring in other Brazilian biomes and forest formations, it is a promising model for developing further dendrochronological research including regional networks of replicated site chronologies, which could facilitate the reconstruction of historical climatic series and predictions of future impacts of climate change in tropical areas.     

Sulfenylation of ENOLASE2 facilitates H2O2-conferred freezing tolerance in Arabidopsis

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A DNA Damage Checkpoint Pathway Coordinates the Division of Dikaryotic Cells in the Ink Cap Mushroom Coprinopsis cinerea

The fungal fruiting body or mushroom is a multicellular structure essential for sexual reproduction. It is composed of dikaryotic cells that contain one haploid nucleus from each mating partner sharing the same cytoplasm without undergoing nuclear fusion. In the mushroom, the pileus bears the hymenium, a layer of cells that includes the specialized basidia in which nuclear fusion, meiosis, and sporulation occur. Coprinopsis cinerea is a well-known model fungus used to study developmental processes associated with the formation of the fruiting body. Here we describe that knocking down the expression of Atr1 and Chk1, two kinases shown to be involved in the response to DNA damage in a number of eukaryotic organisms, dramatically impairs the ability to develop fruiting bodies in C. cinerea, as well as other developmental decisions such as sclerotia formation. These developmental defects correlated with the impairment in silenced strains to sustain an appropriated dikaryotic cell cycle. Dikaryotic cells in which chk1 or atr1 genes were silenced displayed a higher level of asynchronous mitosis and as a consequence aberrant cells carrying an unbalanced dose of nuclei. Since fruiting body initiation is dependent on the balanced mating-type regulator doses present in the dikaryon, we believe that the observed developmental defects were a consequence of the impaired cell cycle in the dikaryon. Our results suggest a connection between the DNA damage response cascade, cell cycle regulation, and developmental processes in this fungus.     

Profiling SARS-CoV-2 HLA-I peptidome reveals T cell epitopes from out-of-frame ORFs

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The Alarmone (p)ppGpp Regulates Primer Extension by Bacterial Primase

Primase is an essential component of the DNA replication machinery, responsible for synthesizing RNA primers that initiate leading and lagging strand DNA synthesis. Bacterial primase activity can be regulated by the starvation-inducible nucleotide (p)ppGpp. This regulation contributes to a timely inhibition of DNA replication upon amino acid starvation in the Gram-positive bacterium Bacillus subtilis. Here, we characterize the effect of (p)ppGpp on B. subtilis DnaG primase activity in vitro. Using a single-nucleotide resolution primase assay, we dissected the effect of ppGpp on the initiation, extension, and fidelity of B. subtilis primase. We found that ppGpp has a mild effect on initiation, but strongly inhibits primer extension and reduces primase processivity, promoting termination of primer extension. High (p)ppGpp concentration, together with low GTP concentration, additively inhibit primase activity. This explains the strong inhibition of replication elongation during starvation which induces high levels of (p)ppGpp and depletion of GTP in B. subtilis. Finally, we found that lowering GTP concentration results in mismatches in primer base pairing that allow priming readthrough, and that ppGpp reduces readthrough to protect priming fidelity. These results highlight the importance of (p)ppGpp in protecting replisome integrity and genome stability in fluctuating nucleotide concentrations upon onset of environmental stress.