Purification and characterization of a novel intracellular α-amylase with a wide variety of substrates hydrolysis and transglycosylation activity from Paenibacillus sp. SSG-1
Contributors: Qingrui Xu, Yu Cao, Xi Li, Lin Liu, Shishang Qin, Yuhao Wang, Yi Cao, Hui Xu, Dairong Qiao
... Intracellular α-amylase was a special glycoside hydrolase in the cytoplasm. We cloned and expressed an intracellular α-amylase, Amy, from Paenibacillus sp. SSG-1. The recombinant enzyme was purified by metal-affinity chromatography, exhibited a molecular mass of 71.7 kDa. Amy exhibited unexpectedly sequence similarity and evolutionary relationships with alpha-glucanotransferase. The docked results of Amy with maltose showed it had similar catalytic residues with α-amylase and glucanotransferase. The substrate specificity experiment showed that Amy could hydrolyze typical substrates into glucose and maltose. It was noteworthy that Amy showed the catalytic capacity of cyclomaltodextrinase and pullulanase. Meanwhile, Amy could transfer sugar molecules and form maltotetraose upon the hydrolysis of substrates. These results indicated that Amy was a novel intracellular α-amylase with distinct catalytic ability characteristics of hydrolyzing glycogen/cyclodextrin/pullulan and transglycosylation. We deduced that Amy may play an important role in utilizing maltooligosaccharides that released from extracellular α-glucan or storage α-glucan (glycogen) in Paenibacillus sp. SSG-1.
Stochastic convergence in per capita CO2 emissions. An approach from nonlinear stationarity analysis
Contributors: María José Presno, Manuel Landajo, Paula Fernández González
... This paper studies stochastic convergence of per capita CO2 emissions in 28 OECD countries for the 1901–2009 period. The analysis is carried out at two aggregation levels: first for the whole set of countries (joint analysis) and then separately for developed and developing states (group analysis). A powerful time series methodology - adapted to a nonlinear framework that allows for quadratic trends with possibly smooth transition between regimes - is applied. This approach provides more robust conclusions in convergence path analysis, enabling (a) robust detection of the presence, and if so, the number of changes in the level and/or slope of the trend of the series; (b) inferences on stationarity of relative per capita CO2 emissions, conditionally on the presence of breaks and smooth transitions between regimes; and (c) estimation of change locations in the convergence paths. Finally, as stochastic convergence is attained when both stationarity around a trend and β-convergence simultaneously hold, the linear approach proposed by Tomljanovich and Vogelsang (2002) is extended in order to allow for more general, quadratic models. Overall, joint analysis finds some evidence of stochastic convergence in per capita CO2 emissions. Some dispersion in terms of β-convergence is detected by the group analysis, particularly among developed countries. This is in accordance with per capita GDP not being the sole determinant of convergence in emissions, with factors like search for more efficient technologies, fossil fuel substitution, innovation, and possibly industry outsourcing, also having a crucial role.
Special issue: Research report - The tract terminations in the temporal lobe: Their location and associated functions
Contributors: Claude J. Bajada, Hamied A. Haroon, Hojjatollah Azadbakht, Geoff J.M. Parker, Matthew A. Lambon Ralph, Lauren L. Cloutman
... Temporal lobe networks are associated with multiple cognitive domains. Despite an upsurge of interest in connectional neuroanatomy, the terminations of the main fibre tracts in the human brain are yet to be mapped. This information is essential given that neurological, neuroanatomical and computational accounts expect neural functions to be strongly shaped by the pattern of white-matter connections. This paper uses a probabilistic tractography approach to identify the main cortical areas that contribute to the major temporal lobe tracts. In order to associate the tract terminations to known functional domains of the temporal lobe, eight automated meta-analyses were performed using the Neurosynth database. Overlaps between the functional regions highlighted by the meta-analyses and the termination maps were identified in order to investigate the functional importance of the tracts of the temporal lobe. The termination maps are made available in the Supplementary Materials of this article for use by researchers in the field.
Production of bioactive recombinant rat soluble receptor for advanced glycation end products (rrsRAGE) in Pichia pastoris
Contributors: Peng Xia, Jin Gao, Wen Guan, Jingjing Li, Xiaolan Yu, Fangyuan Wang, Honglin He, Qing Deng, Liang Zhou, Yunsheng Yuan
... Soluble receptor for advanced glycation end products (sRAGE), a natural inhibitor of RAGE, is considered to be a putative therapeutic molecule for a variety of diseases and a biomarker for certain conditions. To further study the function of sRAGE, recombinant rat sRAGE (rrsRAGE) was expressed and produced in a eukaryotic system. The open reading frame of rat sRAGE was cloned downstream of the methanol-inducible alcohol oxidase promoter of pPICZαA vector, and Pichia pastoris strain X-33 was used as the host strain. The expression of rrsRAGE was achieved by fermentation in a 15-L bioreactor and the resulting fermentation broth was subjected to purification on a cation exchange chromatography column. The purification of rrsRAGE reached 95% after size exclusion chromatography(SEC). The bioactivity of the purified protein was confirmed in a SH-SY5Y cell proliferation assay. The biological function of the purified rrsRAGE protein rat CCl4-induced model was then examined. Treatment with rrsRAGE resulted in significantly lower liver fibrosis and lower serum level of ALT, suggesting that sRAGE prevent liver from injury and fibrosis. In conclusion, we achieved high-efficiency production of bioactive rrsRAGE in Pichia pastoris.
An experience sampling approach to emotion regulation: Situational suppression use and social hierarchy
Contributors: Arman Daniel Catterson, Lameese Eldesouky, Oliver P. John
... Whereas past research has examined the use of emotion regulation strategies in terms of individual differences or responses to experimental manipulations, this research takes a naturalistic and repeated-measures approach to examine suppression use in specific situations. Using an experience sampling design, we find evidence across two samples (total N=215) that (1) there was substantial within-person variation in suppression use, (2) the situational use of suppression was explained by situational differences in extraversion and social hierarchy, and (3) when used in contexts in which people felt they were low in social hierarchy, the negative relationship between suppression and well-being was attenuated. These findings suggest there are contexts in which suppression use may not be maladaptive, and demonstrate the benefits of studying emotion processes in real-life.
Müller glia reactivity follows retinal injury despite the absence of the glial fibrillary acidic protein gene in Xenopus
Contributors: Reyna I. Martinez-De Luna, Ray Y. Ku, Alexandria M. Aruck, Francesca Santiago, Andrea S. Viczian, Diego San Mauro, Michael E. Zuber
... Intermediate filament proteins are structural components of the cellular cytoskeleton with cell-type specific expression and function. Glial fibrillary acidic protein (GFAP) is a type III intermediate filament protein and is up-regulated in glia of the nervous system in response to injury and during neurodegenerative diseases. In the retina, GFAP levels are dramatically increased in Müller glia and are thought to play a role in the extensive structural changes resulting in Müller cell hypertrophy and glial scar formation. In spite of similar changes to the morphology of Xenopus Müller cells following injury, we found that Xenopus lack a gfap gene. Other type III intermediate filament proteins were, however, significantly induced following rod photoreceptor ablation and retinal ganglion cell axotomy. The recently available X. tropicalis and X. laevis genomes indicate a small deletion most likely resulted in the loss of the gfap gene during anuran evolution. Lastly, a survey of representative species from all three extant amphibian orders including the Anura (frogs, toads), Caudata (salamanders, newts), and Gymnophiona (caecilians) suggests that deletion of the gfap locus occurred in the ancestor of all Anura after its divergence from the Caudata ancestor around 290 million years ago. Our results demonstrate that extensive changes in Müller cell morphology following retinal injury do not require GFAP in Xenopus, and other type III intermediate filament proteins may be involved in the gliotic response.
Contributors: Yi Ding, Gabriele Colozza, Kelvin Zhang, Yuki Moriyama, Diego Ploper, Eric A. Sosa, Maria D.J. Benitez, Edward M. De Robertis
... RNA sequencing has allowed high-throughput screening of differential gene expression in many tissues and organisms. Xenopus laevis is a classical embryological and cell-free extract model system, but its genomic sequence had been lacking due to difficulties arising from allotetraploidy. There is currently much excitement surrounding the release of the completed X. laevis genome (version 9.1) by the Joint Genome Institute (JGI), which provides a platform for genome-wide studies. Here we present a deep RNA-seq dataset of transcripts expressed in dorsal and ventral lips of the early Xenopus gastrula embryo using the new genomic information, which was further annotated by blast searches against the human proteome. Overall, our findings confirm previous results from differential screenings using other methods that uncovered classical dorsal genes such as Chordin, Noggin and Cerberus, as well as ventral genes such as Sizzled, Ventx, Wnt8 and Bambi. Complete transcriptome-wide tables of mRNAs suitable for data mining are presented, which include many novel dorsal- and ventral-specific genes. RNA-seq was very quantitative and reproducible, and allowed us to define dorsal and ventral signatures useful for gene set expression analyses (GSEA). As an example of a new gene, we present here data on an organizer-specific secreted protein tyrosine kinase known as Pkdcc (protein kinase domain containing, cytoplasmic) or Vlk (vertebrate lonesome kinase). Overexpression experiments indicate that Pkdcc can act as a negative regulator of Wnt/ β-catenin signaling independently of its kinase activity. We conclude that RNA-Seq in combination with the X. laevis complete genome now available provides a powerful tool for unraveling cell-cell signaling pathways during embryonic induction.
Pancreatic proteome profiling of type 1 diabetic mouse: Differential expression of proteins involved in exocrine function, stress response, growth, apoptosis and metabolism
Contributors: Bulbul Chakravarti, Chheten Sherpa, Devasrie Bose, Kakoli Paul Chowdhury, Kavita Khadar, Y. Clare Zhang, Deb N. Chakravarti
... Type 1 diabetes (T1D) is a chronic autoimmune disease in which the pancreatic β-cells fail to produce insulin. In addition to such change in the endocrine function, the exocrine function of the pancreas is altered as well. To understand the molecular basis of the changes in both endocrine and exocrine pancreatic functions due to T1D, the proteome profile of the pancreas of control and diabetic mouse was compared using two dimensional gel electrophoresis (2D-GE) and the differentially expressed proteins identified by electrospray ionization liquid chromatography-tandem mass spectrometry (ESI-LC-MS/MS). Among several hundred protein spots analyzed, the expression levels of 27 protein spots were found to be up-regulated while that of 16 protein spots were down-regulated due to T1D. We were able to identify 23 up-regulated and 9 down-regulated protein spots and classified them by bioinformatic analysis into different functional categories: (i) exocrine enzymes (or their precursors) involved in the metabolism of proteins, lipids, and carbohydrates; (ii) chaperone/stress response; and (iii) growth, apoptosis, amino acid metabolism or energy metabolism. Several proteins were found to be present in multiple forms, possibly resulting from proteolysis and/or post-translational modifications. Succinate dehydrogenase [ubiquinone] flavoprotein subunit, which is the major catalytic subunit of succinate dehydrogenase (SDH), was found to be one of the proteins whose expression was increased in T1D mouse pancreata. Since altered expression of a protein can modify its functional activity, we tested and observed that the activity of SDH, a key metabolic enzyme, was increased in the T1D mouse pancreata as well. The potential role of the altered expression of different proteins in T1D associated pathology in mouse is discussed.
Clinical Study - Validation of the Spine Oncology Study Group—Outcomes Questionnaire to assess quality of life in patients with metastatic spine disease
Contributors: Stein J. Janssen, Teun Teunis, Eva van Dijk, Marco L. Ferrone, John H. Shin, Francis Hornicek, Joseph H. Schwab
... General questionnaires are often used to assess quality of life in patients with spine metastases, although a disease-specific survey did not exist until recently. The Spine Oncology Study Group has developed an outcomes questionnaire (SOSG-OQ) to measure quality of life in these patients. However, a scoring system was not developed, and the questionnaire was not validated in a group of patients, nor was it compared with other general quality of life questionnaires such as the EuroQol 5 Dimensions (EQ-5D) questionnaire.
Overexpression of the immediate-early genes Egr1, Egr2, and Egr3 in two strains of rodents susceptible to audiogenic seizures
Contributors: D. López-López, R. Gómez-Nieto, M.J. Herrero-Turrión, N. García-Cairasco, D. Sánchez-Benito, M.D. Ludeña, D.E. López
... Genetic animal models of epilepsy are an important tool for further understanding the basic cellular mechanisms underlying epileptogenesis and for developing novel antiepileptic drugs. We conducted a comparative study of gene expression in the inferior colliculus, a nucleus that triggers audiogenic seizures, using two animal models, the Wistar audiogenic rat (WAR) and the genetic audiogenic seizure hamster (GASH:Sal). For this purpose, both models were exposed to high intensity auditory stimulation, and 60min later, the inferior colliculi were collected. As controls, intact Wistar rats and Syrian hamsters were subjected to stimulation and tissue preparation protocols identical to those performed on the experimental animals. Ribonucleic acid was isolated, and microarray analysis comparing the stimulated Wistar and WAR rats showed that the genomic profile of these animals displayed significant (fold change, |FC|≥2.0 and p<0.05) upregulation of 38 genes and downregulation of 47 genes. Comparison of gene expression profiles between stimulated control hamsters and stimulated GASH:Sal revealed the upregulation of 10 genes and the downregulation of 5 genes.