Miracle-dz/UniFAR-FTUs huggingface dataset:
Engineering Lactococcus lactis for the scaffold protein-mediated surface display of recombinant enzymes

Multi-enzyme complexes are responsible for the synthesis of a number of biochemical compounds and the degradation of complex polymers. An example of the latter is the degradation of cellulose by enzyme complexes termed “cellulosomes” which are produced by several bacteria of the class Clostridia. The basic structure of a cellulosome comprises a central scaffold protein, which associates with a multitude of cellulases via cohesin-dockerin interactions. The advent of cellulose utilization as feedstock for producing biofuels has garnered much interest towards designing custom-tailored recombinant cellulosomes and expressing them in microbes of interest. The metabolic diversity among bacteria also make this approach an appealing strategy for bestowing cellulolytic capabilities upon organisms which produce non-biofuel commodity chemicals such as lactic acid, succinic acid, acetone, amino acids, food additives and carotenoids. In addition, the display of recombinant multi-enzyme complexes in bacteria can yield novel insights into the mechanisms and parameters affecting their secretion, assembly and function. The industrially relevant lactic acid bacterium, Lactococcus lactis, is a model organism for the secretion and display of recombinant proteins, and the numerous biological techniques available for its manipulation make this organism particularly appealing for such a task. In this thesis, I present my work describing the incremental steps taken towards the surface display of custom-tailored multi-enzyme complexes on the surface of L. lactis. Chapter 1 describes the proof of concept for this project, including the choice of promoters, secretion signal peptide, and reporter enzymes. It also discusses the major bottlenecks observed based on the organism’s physiology. Chapter 2 describes the engineering of scaffold chimeras with cohesins of different specificity and the display of two enzymes on such scaffolds. I also investigated the catalytic profiles of the resulting complexes when enzymes were simultaneously or sequentially bound to the displayed scaffold. Finally, chapter 3 describes the optimization of the type 2 dockerin-cohesin interaction by the inclusion of the CipA “X” module, as well as the engineering of enzyme complexes with novel architectures by use of secondary “adapter” scaffolds and the subsequent assembly of multi-scaffold complexes. Also investigated is the potential of using a dual-plasmid system for the full in vivo assembly of such complexes without the exogenous addition of components.
Assembly and composition of the cellular actin cortex

The actin cortex is a thin layer of F-actin, actin regulating proteins, and myosin under the cell membrane. It has a fundamental role in cell morphogenesis, motility, mitosis, cytokinesis, and cell shape maintenance. Yet, how the cell cortex forms as well as the protein composition of the cortex remain unknown. In this study using M2 melanoma cell blebs and metaphase HeLa cells I describe proteins needed for cortex reassembly and reveal the protein composition of the actin cortex. Blebs represent a natural situation in which the membrane is transiently devoid of an actin cortex. Thus the formation of a new actin cortex in the bleb can be studied. Blebs also provide a cell fraction enriched in F-actin cortex, making them an ideal model to study cortex proteomics. After having shown that no nucleation independent mechanism participate in cortex reassembly in membrane blebs, I discovered by localisation and silencing studies, that formin Diaph1 and the Arp2/3 complex are needed for the cortex nucleation, as both localised to the cortex and depletion of either led to cortical defects. Simultaneous depletion of Diaph1 and the Arp2/3 complex led to disappearance of the cellular cortex, suggesting that together they provide the majority of the F-actin in the cortex. Cortex composition studies of blebs isolated from blebbing cells revealed the presence of many actin binding proteins at the cortex. Localisation and depletion studies revealed that capping, and effective actin turnover are essential for cortical stability and that flightless-I may regulate cortex nucleation together with Rho GTPases. These findings have important implications for our understanding of cortical properties, which are not only important for cell morphogenesis and shape maintenance, but facilitate tumour growth and dissemination.
Biochemical basis of human disease-causing actin mutations

Actin isoform specific mutations have been identified as causes for various human diseases. These include twelve missense mutations in γ-nonmuscle actin leading to early onset autosomal dominate nonsyndromic hearing loss and twenty two missense mutations in α-smooth muscle actin leading to thoracic aortic aneurysms and dissections (TAAD). The molecular mechanisms leading to these human pathologies are mainly unknown, principally due to the inability to isolate pure mutant γ-nonmuscle actin and αsmooth muscle actin in quantities required for biochemical analysis. To begin to address these problems, I have individually expressed the human nonmuscle actin isoforms, βand γnonmuscle actin, in a baculovirus expression system and characterized their biochemical properties. Surprisingly, despite a conserved amino acid difference of only 4 residues at or near the N-terminus, Ca-γ-actin exhibits slower monomeric and filamentous biochemical properties than β-actin. In the Mg-form, the difference between the two is smaller. Mixing experiments with Ca-actins reveal the two will readily copolymerize. Calcium bound in the high affinity binding site of γ-actin may cause a selective energy barrier relative to β-actin that retards the equilibration between Gand Fmonomer conformations resulting in a slower polymerizing actin with greater filament stability. This difference may be particularly important in sites such as the γ-actin-rich cochlear hair cell stereocilium where local mM calcium concentrations may exist. In hair cells γ-nonmuscle actin seems to play a central role in stereocilia maintenance. To determine how the deafness causing D51N-γ-mutant actin mutation leads to deafness, I expressed and characterized it in the γ-actin background. The D51N mutation, lethal when cloned into yeast, displayed decreased filament stability and polymerization kinetics of an actin more dynamic than γ-actin. This result suggests that the hearing effects of the γ-actin mutations on the hearing apparatus are not simply caused by an inability to polymerize. The observed increased polymerization rates and
Genetic diversity of Vetiver clones (Chrysopogon zizanioides and Chrysopogon nigritana) available in South Africa based on sequencing analyses and anatomical structure

Vetiver grass or Chrysopogon zizanioides (L.) Roberty (1960) is a sterile grass which can regenerate vegetatively from clumps of the rootstock. This, as well as its vigorous and deep root system and flood tolerance makes it an ideal candidate for the use in soil remediation and erosion control. In South Africa, Hydromulch (Pty) Ltd. is part of the landscape, soil reclamation and erosion control industry. The company uses vetiver grass on a wide scale
Cholesterol and fatty acid content in meat of turkeys fed diets with sunflower, linseed or fish oil

Male and female turkeys of BUT Big 6 hybrid were fed either the control diet (C) or the diet enriched with sunflower oil (S), linseed oil (L) and fish oil (F), respectively. Eight birds from each group within each sex were slaughtered at 56 days and breast meat (BM) and thigh meat (TM) was separated. Total lipid (TL) was extracted from separated tissues using hexane/2-propanol mixture, and total cholesterol (by liquid chromatography) and 15 fatty acids (by gas chromatography) were determined in the extract. TL content did not differ (P > 0.05) between dietary treatments either in BM (0.73-0.78%) or in TM of males (2.22-2.63%). However, S-females deposited more (P 0.05) from corresponding tissues of F-groups (0.9 in BM and 1.4 in TM). The diet with linseed oil had also the same (P > 0.05) effect as F-diet on deposition of arachidonic acid in both BM (22-25 mg/100 g) and TM (33-34 mg/ 100 g) of both sexes. On the other hand, PUFA n-6/n-3 ratio (6-7 in BM, 16-17 in TM) and arachidonic acid content (54-57 and 69-76 mg/100 g in BM and TM, respectively) in both tissues of both sexes was the highest (P 0.05) percentage in BM and nearly twice as high (P < 0.05) percentage in TM of total n-3 PUFA in both L-groups in comparison with F-ones, the equivalent of very long chain n-3 PUFA, VLCE (VLCE = 0.15 x α-linolenic acid + [eicosapentaenoic acid + docosahexaenoic acid]) was both in BM and in TM of L-turkeys substantially lower (P < 0.05) than in F-birds (66-68 vs. 114-118 and 145-158 vs. 245-262 mg/100 g, respectively). It was concluded, that linseed oil in an amount of 5% of dietary crude fat was not sufficient to substitute fish oil as regards content of VLCE in turkey meat, but had the same (n-6/n-3 PUFA ratio, arachidonic acid content) or a better (cholesterol content) effect on some other parameters of nutritive value of turkey BM and TM.
Estimation of temporal variations in historical fire frequency from time-since-fire map data

This paper describes an improved statistical methodology for estimating historical forest fire frequencies from time-since-fire map data. Of particular interest is the question of estimating fire frequency in temporally distinct epochs. Unlike previous graphical and ad hoc methods, the new methodology is firmly grounded within the established statistical paradigm of likelihood inference. In addition, in contrast to earlier methods, it takes into account the fact that surviving stands which originated in earlier epochs, have been subject to different hazards of burning as they have lived through more recent epochs. Neglecting this fact leads to overestimates of the fire frequency in the more recent epochs. Procedures for obtaining maximum likelihood estimates and likelihood ratio confidence intervals for the fire frequency in each epoch are developed, along with a likelihood ratio test to assess whether the fire frequencies in distinct epochs were significantly different. This test is not strictly valid when the change points separating epochs are suggested from exploratory data analysis; rather it is developed assumingthat change points are determined independently of the data. This distinction, which is necessary to avoid selection bias, has not been noted before. The fact that forest fire is a contagious process, with consequent spatial autocorrelation in time-since-fire observations, is taken into account through the use of an overdispersed model, with associated quasilikelihood function. This aspect has been ignored, by and large, in previous analyses which have implicitly assumed independence of observations. An example is given in which the method is applied to published fire history data from the Kananaskis Valley. The results indicate that the previous an alysis overestimated the fire frequency for the most recent epoch. The conclusions of other studies concerning temporal changes in fire frequency may need to be reconsidered. FOR. Sc•. 44(3):465-475. Additional
Dealing with complexity of new phenotypes in modern dairy cattle breeding

April 2020, Vol. 10, No. 2 This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons. org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com © Seidel, Krattenmacher, and Thaller Feature Article Dealing with complexity of new phenotypes in modern dairy cattle breeding
Coping with stress: Anaerobic respiratory and oxidative stress tolerance mechanisms are critical for Neisseria gonorrhoeae biofilm formation

Many illnesses and infections are exacerbated and/or caused by biofilms. Neisseria gonorrhoeae, the etiologic agent of gonorrhea, is frequently asymptomatic in women, which can lead to persistent infection. Persistent infection can result in pelvic inflammatory disease, tubo-ovarian abscesses, infertility, and ectopic pregnancy. N. gonorrhoeae has been shown to form biofilms over glass, primary and immortalized cervical cells, and during natural cervical infection. Asymptomatic infection occurs in only 1% of infected males, and the infection site is subject to periodic rapid fluid flow, which may limit biofilm formation. Thus, biofilm formation may specifically play an important role in the infection of women and could contribute to the infrequent occurrence of symptoms. Prior to work presented in this dissertation, little was known about biofilm formation by N. gonorrhoeae. Therefore, we elected to compare the transcriptional profiles of biofilms to their planktonic counterparts, and to identify genetic pathways involved in biofilm formation and maintenance. We found that 3.8% of the genome was differentially regulated, and that genes involved in anaerobic metabolism and oxidative stress tolerance were up-regulated in biofilm, while genes involved in aerobic metabolism were down-regulated. We determined that expression of aniA, ccp, and norB is required for robust biofilm formation over glass and human cervical cells, and anaerobic respiration occurs in the substratum of gonococcal biofilms. Disruption of the norB gene resulted in severe attenuation of biofilm formation. We determined that the accumulation of nitric oxide (NO) contributes to the phenotype of a norB mutant and can retard biofilm formation when present at sublethal concentrations. However, higher concentrations of NO can enhance biofilm formation in the absence of nitrite. NO enhances biofilm formation in an aniA mutant, but cannot completely restore
Localisation chromosomique et expression tissulaire des gènes amplifiés codant les estérases impliquées dans la résistance aux insecticides organophosphorés chez "Culex pipiens"

Chez le moustique culex pipiens, la resistance aux insecticides orgosphosphores peut etre due a une augmentation de la detoxication par des esterases qui sont surproduites grace a l'amplification des genes qui les codent. Nous montrons, en utilisant des techniques d'hybridation in situ et d'immunohistochimie, que les genes amplifies sont sur un meme chromosome et nous identifions les tissus dans lesquels ils s'expriment. Les esterases de detoxication n'existent que dans certains tissus de l'animal et la nature de ces tissus varie selon les souches. Il semble que la distribution tissulaire des esterases surproduites soit un facteur capital dans l'acquisition d'un taux de resistance eleve
Echanges d'azote ureique et ammoniacal a travers la paroi du rumen de mouton : cinetique, bilan journalier et mecanismes de regulation

Pour ameliorer la connaissance des processus de transfert d'uree et d'ammoniaque a travers la paroi du rumen de moutons, une technique permettant de mesurer de facon directe et continue les flux de nutriments a travers l'epithelium du rumen a ete developpee. Les moutons ont ete prepares chirurgicalement de la facon suivante: implantation d'un catheter arteriel et d'un catheter dans chacune des veines du rumen; pose de sondes debitmetriques sur les arteres ruminales; canulation du rumen. L'utilisation de catheters en silicone a l'interieur des veines ruminales, et la mesure du flux sanguin par debitmetrie ultrasonique, permettent d'obtenir des mesures fiables pour des periodes allant jusqu'a 6 mois. Le debit sanguin ruminal augmente largement lors de l'ingestion d'un repas, cette augmentation est d'autant plus importante que la quantite de matiere seche ingeree augmente. Les flux de metabolites a travers la paroi du rumen ont ete etudies sur des moutons adultes, recevant un repas de 500 g de foin de dactyle toutes les 12 h. L'absorption d'ammoniaque est intense pendant le repas, alors que 3 h apres elle est redescendue a son niveau preprandial. Par contre le transfert net d'uree augmente des la fin du repas, et atteint son maximum 5 h apres, au moment ou la concentration en ammoniaque intraruminale est au plus bas. Les flux nets d'ammoniaque et d'uree a travers la paroi du rumen ont ete estimes respectivement a 3. 76 et 2. 10 g n/j. La concentration en ammoniaque non ionise dans le rumen et le debit sanguin ruminal paraissent etre les principaux facteurs de la regulation a court terme de l'absorption d'ammoniaque. Le flux transepithelial d'uree est facilite par les ureases bacteriennes intraruminales. L'insufflation de co#2 dans le rumen stimule la diffusion de l'uree a travers la paroi. Le flux d'uree est negativement relie a l'absorption d'ammoniaque
Analysis of Azurin Protein from Different Pseudomonas sp Using Proteomic Tools

Abstract: Azurin is copper-containing protein called cupredoxin protein. This protein mainly involve in electron-transfer. In the present study, azurin proteins from different Pseudomonas species were analysed for their primary, secondary structure, physiochemical analysis and phylogenetic relation. The present study indicated that amino acid composition, molecular weight varies among the azurin protein. Alanine is the most common and abundant amino acid. The phylogenetic analysis showed that the azurin protein of various species of Pseudomonas had a common ancestor, closely related and the sequences are highly conserved. Key words: Copper protein, electron transfer, protein sequence analysis, bacteria, bioinformatics tools. 1. Introduction Azurin is one of a class of copper-containing proteins called blue-copper proteins, so called because of their striking blue colour. Azurin from Pseudomonas species are involved in an electron-transfer hence, it is called cupredoxin. However, it physiological function is not precisely known [1, 2] (Fig. 1).
Family Nurse Practitioner Student Competencies in Integrated Psychiatric Healthcare

Title : Practicum to Practice: Family Nurse Practitioner Student Competencies in Integrated Psychiatric Mental Healthcare. Background: Major Depressive Disorder (MDD) is a widespread and costly diagnosis that affects nearly 16 million people annually and costs the United States nearly $83 Billion. Primary care providers such as Family Nurse Practitioners (FNP) play a crucial role in screening for health problems, preventing, and managing illness. Opportunities exist to integrate psychiatric mental healthcare into FNP curriculum and clinical practicums to better prepare providers to identify and screen for depression. Purpose: The purpose of this evidenced-based practice project is to evaluate FNP student competencies in screening for depression after completing an integrated family/psychiatric mental health curriculum and clinical practicum. Program Plan Process: An integrated family/psychiatric mental health curriculum and clinical practicum was developed to educate FNP students to identify and screen for depressed patients. FNP students will demonstrate depression screening competencies utilizing a simulated clinical encounter in a simulation center. Four students received the integrated intervention (intervention group) and four students did not receive the intervention (non-intervention group). Results: The integrated clinical curriculum and practicum students showed less variability in performance and demonstrated more consistency with treatment compared with the non-intervention group.
Biology and regulation of carcass composition

This chapter provides an overview of the generally accepted understanding of the biology and regulation of carcass composition. More specifically, patterns of growth are briefly described and an overview of the biology of carcass tissue growth and development is provided. Regulation of compositional characteristics by nutrition, genotype and metabolic modifiers is also reviewed. Finally, future trends in research relating to the regulation of carcass composition and meat production are discussed.
Validation of a semiquantitative food‐frequency questionnaire measuring dietary vitamin K intake in elderly

The study objective was to validate a semiquantitative food frequency questionnaire (FFQ) designed to measure dietary vitamin K (VK) intake specifically. A 50‐item FFQ was interviewer‐administered and compared with data previously obtained from 5‐d food records (FR). Thirty‐nine community‐dwelling healthy men and women aged 65 to 85 years were recruited from the Montréal metropolitan area. Absolute and relative agreements between methods were assessed. Vitamin K intakes measured by the VK‐FFQ (mean ± SD; 222 ± 186 μg/d) were 2.26 times (95%CI = 1.90, 2.67) higher than those obtained by FRs (135 ± 153 μg/d; P< 0.001). However, cross‐classification showed that 72% of participants were correctly classified into thirds and only 8% were grossly classified. Kappa value (κ=0.60) and correlation coefficient (r=0.83) also indicated a good relative agreement. In light of these results, the VK‐FFQ developed is a valid tool for ranking individuals according to their VK intakes. The poor absolute agreement likely results from the inability for FRs to adequately measure the usual intake of episodically consumed foods. The VK‐FFQ will be useful in large‐scale, population‐based research on VK and diseases as well as in clinical practice, especially that focusing on anticoagulant therapy. This research was supported by the Canadian Institutes of Health Research.
THE OVARY OF Mustelus schmitti (CHONDRICHTHYES, TRIAKIDAE): MICROANATOMY AND DEVELOPMENT THROUGH THE REPRODUCTIVE CYCLE

This work is the first approach to the ovary microanatomy of Mustelus sch mitti (gatuso) through their reproductive cycle, including the follicular p opulation and the characteristics of the stroma. Three macroscopic stage s of sexual maturity were considered. The ovaries of im mature females show only the early phases of foliculogenesis. In young anima ls there are the first indications of follicular atresia and in mature ones all f ollicular stages a re present as well as corpora lutea. The ovogonias a re present through the entire reproductive cycle.
Functions of retrograde axonal transport.

: Retrograde axonal transport conveys materials from axon to cell body. One function of this process is recycling of materials originally transported from cell body to axon. In motoneurons, 50% of fast-transported protein is returned. Reversal probably occurs mainly at nerve terminals and, for labeled proteins, is nonselective. Proteolysis is not required, although changes in tertiary protein structure may occur with a repackaging of molecules in organelles different from those in which they were anterograde-transported. A second function is transfer of information about axonal status and terminal environment. Premature reversal of transport adjacent to an axon injury may be a component of a signal that initiates cell body chromatolysis. Transport of target cell-derived molecules with trophic effects on the cell body is exemplified by nerve growth factor transport in neurons dependent on it, and is probably a widespread phenomenon in the developing nervous system. Disorders in retrograde transport or reversal occur in some experimental neuropathies, and certain viruses, as well as tetanus toxin, may gain access to the central nervous system by this route.
Plant Growth Regulators from Kenyan Plant, Psiadia punctulata

Compounds having plant growth regulatory effect were found in Kenyan Psiadia punctulata, and two flavones,5-hydroxy-7,2',3',4',5'-pentamethoxyflavone(1) and 5,3'-dihydroxy-7,2',4',5'-tetramethoxyflavone(2) were isolated and identified as the activ constituents.