Articles from MDPI


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  • Itkonen, Jaakko; Annala, Ada; Tavakoli, Shirin; Arango-Gonzalez, Blanca; Ueffing, Marius; Toropainen, Elisa; Ruponen, Marika; Casteleijn, Marco G; Urtti, Arto (Multidisciplinary Digital Publishing Institute, 2020)
    Ciliary neurotrophic factor (CNTF) is one of the most studied neuroprotective agents with acknowledged potential in treating diseases of the posterior eye segment. Although its efficacy and mechanisms of action in the retina have been studied extensively, it is still not comprehensively understood which retinal cells mediate the therapeutic effects of CNTF. As with therapeutic proteins in general, it is poorly elucidated whether exogenous CNTF administered into the vitreous can enter and distribute into the retina and hence reach potentially responsive target cells. Here, we have characterized our purified recombinant human CNTF (rhCNTF), studied the protein’s in vitro bioactivity in a cell-based assay, and evaluated the thermodynamic and oligomeric status of the protein during storage. Biological activity of rhCNTF was further evaluated in vivo in an animal model of retinal degeneration. The retinal penetration and distribution of rhCNTF after 24 h was studied utilizing two ex vivo retina models. Based on our characterization findings, our rhCNTF is correctly folded and biologically active. Moreover, based on initial screening and subsequent follow-up, we identified two buffers in which rhCNTF retains its stability during storage. Whereas rhCNTF did not show photoreceptor preservative effect or improve the function of photoreceptors in vivo, this could possibly be due to the used disease model or the short duration of action with a single intravitreal injection of rhCNTF. On the other hand, the lack of in vivo efficacy was shown to not be due to distribution limitations; permeation into the retina was observed in both retinal explant models as in 24 h rhCNTF penetrated the inner limiting membrane, and being mostly observed in the ganglion cell layer, distributed to different layers of the neural retina. As rhCNTF can reach deeper retinal layers, in general, having direct effects on resident CNTF-responsive target cells is plausible.
  • Salih, Enass Y. A.; Julkunen-Tiitto, Riitta; Luukkanen, Olavi; Sipi, Marketta; Fahmi, Mustafa K. M.; Fyhrquist, Pia Johanna (Multidisciplinary Digital Publishing Institute, 2020)
    In Sudanese traditional medicine, decoctions of the stem bark of <i>Anogeissus leiocarpa</i> are used for the treatment of tuberculosis (TB). However, this plant has not been investigated before for its antimycobacterial effects. Our screening results show, for the first time, that many extracts of various parts of <i>A. leiocarpa</i> exhibit growth inhibitory activity against <i>Mycobacterium smegmatis</i>. Minimum inhibitory concentration (MIC) values ranged between 625 and 5000 &micro;g/mL, with an ethyl acetate extract of the root showing the lowest MIC value. The good antimycobacterial effects of the root part could be due to its high concentration of ellagic acid derivatives, ellagitannins, and flavonoids. Thin layer chromatography (TLC) fractionation resulted in some fractions with better activity than the starting point crude methanol extract (MIC 2500 &micro;g/mL). Those fractions with the lowest MIC values contained a high number of antioxidant compounds. Fractions <b>3</b> and <b>4</b> (MIC 1500 and 1000 &micro;g/mL, respectively) contained high concentrations of di-methyl ellagic acid ([M-H]<sup> &minus;</sup> 329.0318). Fraction <b>6</b> (MIC 2000 &micro;g/mL) contained a lower concentration of di-methyl ellagic acid and was not as growth inhibitory as fractions <b>3</b> and <b>4</b>. Moreover, in fraction <b>3</b>, an acetylated ellagic acid derivative ([M-H]<sup> &minus;</sup> 343.0477) and di-methyl-ellagic acid xyloside ([M-H]<sup>&minus;</sup> 461.0739) were tentatively characterized. Di-methyl ellagic acid xyloside was also present in fraction <b>4</b> and could strongly contribute to the antimycobacterial effect of this fraction. Additionally, protocatechuic acid ([M-H]-at m/z 153.0196) was present in fraction <b>4</b>. Our antimycobacterial results obtained from this research justify the use of <i>A. leiocarpa</i> in Sudanese folk medicine against cough related to TB. Roots, stem bark, and leaves of <i>A. leiocarpa</i> are sources for new potent anti-TB drug lead compounds.
  • Saarinen, Niila V. V.; Lehtonen, Jussi; Veijola, Riitta; Lempainen, Johanna; Knip, Mikael; Hyöty, Heikki; Laitinen, Olli H.; Hytönen, Vesa P. (Multidisciplinary Digital Publishing Institute, 2020)
    Immunological assays detecting antibodies against enteroviruses typically use a single enterovirus serotype as antigen. This limits the ability of such assays to detect antibodies against different enterovirus types and to detect possible type-specific variation in antibody responses. We set out to develop a multiplexed assay for simultaneous detection of antibodies against multiple enterovirus and rhinovirus types encompassing all human infecting species. Seven recombinant VP1 proteins from enteroviruses EV-A to EV-D and rhinoviruses RV-A to RV-C species were produced. Using Meso Scale Diagnostics U-PLEX platform we were able to study antibody reactions against these proteins as well as non-structural enterovirus proteins in a single well with 140 human serum samples. Adults had on average 33-fold stronger antibody responses to these antigens (<i>p</i> &lt; 10<sup>&minus;11</sup>) compared to children, but children had less cross-reactivity between different enterovirus types. The results suggest that this new high-throughput assay offers clear benefits in the evaluation of humoral enterovirus immunity in children, giving more exact information than assays that are based on a single enterovirus type as antigen.
  • Wolff, Lili-Ann (Multidisciplinary Digital Publishing Institute, 2020)
    Humans have always lived in eras of more or less obvious crises and risks. When Ulrich Beck wrote about the risk society in 1986, he talked about risks as invisibility lacking spatial and temporal boundaries. The environmental risks of the modern society may often appear diffuse, even if, for example, the climate change dilemma has progressively become noticeable. However, this year, people on Earth have had to face a most obvious risk. The effects of Covid-19 have reached such proportions that the human world probably will never be the same again. However, the extent of jeopardies is not similar for all world inhabitants, neither are the tools to handle the risks. To face the threat and learn from it, humans need to change manners on many levels and in many social and physical areas. Some of the main questions to reflect on and discuss in this feature issue of <i>Sustainability</i> are: What will the most urgent role of sustainability education be now and in the future? What kind of teaching, learning and educational policies are most relevant? What issues are most crucial in sustainability education research?
  • Ianevski, Aleksandr; Kulesskiy, Evgeny; Krpina, Klara; Lou, Guofeng; Aman, Yahyah; Bugai, Andrii; Aasumets, Koit; Akimov, Yevhen; Bulanova, Daria; Gildemann, Kiira; Arutyunyan, Albert F.; Susova, Olga Yu.; Zhuze, Alexei L.; Ji, Ping; Wang, Wei; Holien, Toril; Bugge, Marit; Zusinaite, Eva; Oksenych, Valentyn; Lysvand, Hilde; Gerhold, Joachim M.; Bjørås, Magnar; Johansen, Pål; Waage, Anders; Heckman, Caroline A.; Fang, Evandro F.; Kainov, Denis E. (Multidisciplinary Digital Publishing Institute, 2020)
    Background: The evidence that pan-Bcl-2 or Bcl-xL-specific inhibitors prematurely kill virus-infected or RNA/DNA-transfected cells provides rationale for investigating these apoptotic inducers further. We hypothesized that not only invasive RNA or DNA (biological factors) but also DNA/RNA-damaging chemical or physical factors could trigger apoptosis that have been sensitized with pan-Bcl-2 or Bcl-xL-specific agents; Methods: We tested chemical and physical factors plus Bcl-xL-specific inhibitor A-1155463 in cells of various origins and the small roundworms (<i>C. elegans</i>); Results: We show that combination of a A-1155463 along with a DNA-damaging agent, 4-nitroquinoline-1-oxide (4NQO), prematurely kills cells of various origins as well as <i>C. elegans</i>. The synergistic effect is p53-dependent and associated with the release of Bad and Bax from Bcl-xL, which trigger mitochondrial outer membrane permeabilization. Furthermore, we found that combining Bcl-xL-specific inhibitors with various chemical compounds or physical insults also induced cell death; Conclusions: Thus, we were able to identify several biological, chemical and physical triggers of the evolutionarily conserved Bcl-xL-mediated apoptotic pathway, shedding light on strategies and targets for novel drug development.
  • Heinonen, Annu; Jakobsson, Maija; Kiviharju, Mari; Virtanen, Seppo; Aro, Karoliina; Kyrgiou, Maria; Nieminen, Pekka; Kalliala, Ilkka (Multidisciplinary Digital Publishing Institute, 2020)
    Colposcopy is often used in follow-up after treatment for cervical intraepithelial neoplasia (CIN) despite its marked inter-observer variability and low sensitivity. Our objective was to assess the role of colposcopy in post-treatment follow-up in comparison to hrHPV (high-risk human papillomavirus) testing, cytology, and cone margin status. Altogether, 419 women treated for histological high-grade lesion (HSIL) with large loop excision of the transformation zone (LLETZ) attended colposcopy with cytology and hrHPV test at six months. Follow-up for recurrence of HSIL continued for 24 months. Colposcopy was considered positive if colposcopic impression was recorded as high grade and cytology if HSIL, ASC-H (atypical squamous cells, cannot exclude HSIL), or AGC-FN (atypical glandular cells, favor neoplasia) were present. Overall, 10 (10/419, 2.4%) recurrent HSIL cases were detected, 5 at 6 months and 5 at 12 months. Colposcopic impression was recorded at 407/419 6-month visits and was positive for 11/407 (2.7%). None of them had recurrent lesions, resulting in 0% sensitivity and 97% specificity for colposcopy. Sensitivity for the hrHPV test at 6 months was 100% and specificity 85%, for cytology 40% and 99%, and for margin status at treatment 60% and 82%, respectively. While the hrHPV test is highly sensitive in predicting recurrence after local treatment for CIN, colposcopy in an unselected population is not useful in follow-up after treatment of CIN.
  • Heikkurinen, Pasi; Ruuska, Toni; Valtonen, Anu; Rantala, Outi (Multidisciplinary Digital Publishing Institute, 2020)
    The Special Issue on &lsquo;After the Anthropocene: Time and Mobility&rsquo; is published. It discusses the geological time to follow the human-dominated epoch and ways to move there. In addition to this editorial, a total of five articles are published in the issue. The articles engage with a variety of social science disciplines&mdash;ranging from economics and sociology to philosophy and political science&mdash;and connect to the natural science insights on the Anthropocene. The issue calls for going beyond anthropocentrism in sustainability theory and practice in order to exit the Anthropocene with applications and insights in the contexts of politics (Ruuska et al., 2020), energy (Mohorčich, 2020), tourism (Rantala et al., 2020), food (Mazac and Tuomisto, 2020) and management (K&uuml;pers, 2020). We hope that you will find this Special Issue interesting and helpful in contributing to sustainable change.
  • Feola, Sara; Chiaro, Jacopo; Martins, Beatriz; Cerullo, Vincenzo (Multidisciplinary Digital Publishing Institute, 2020)
    According to the latest available data, cancer is the second leading cause of death, highlighting the need for novel cancer therapeutic approaches. In this context, immunotherapy is emerging as a reliable first-line treatment for many cancers, particularly metastatic melanoma. Indeed, cancer immunotherapy has attracted great interest following the recent clinical approval of antibodies targeting immune checkpoint molecules, such as PD-1, PD-L1, and CTLA-4, that release the brakes of the immune system, thus reviving a field otherwise poorly explored. Cancer immunotherapy mainly relies on the generation and stimulation of cytotoxic CD8 T lymphocytes (CTLs) within the tumor microenvironment (TME), priming T cells and establishing efficient and durable anti-tumor immunity. Therefore, there is a clear need to define and identify immunogenic T cell epitopes to use in therapeutic cancer vaccines. Naturally presented antigens in the human leucocyte antigen-1 (HLA-I) complex on the tumor surface are the main protagonists in evocating a specific anti-tumor CD8+ T cell response. However, the methodologies for their identification have been a major bottleneck for their reliable characterization. Consequently, the field of antigen discovery has yet to improve. The current review is intended to define what are today known as tumor antigens, with a main focus on CTL antigenic peptides. We also review the techniques developed and employed to date for antigen discovery, exploring both the direct elution of HLA-I peptides and the in silico prediction of epitopes. Finally, the last part of the review analyses the future challenges and direction of the antigen discovery field.
  • Pihlström, Sami (Multidisciplinary Digital Publishing Institute, 2020)
    This paper proposes to reconsider agnosticism by taking a step onto a meta-level, investigating agnosticism not as an epistemic stance regarding the choice between theism and atheism, but as a stance toward the question concerning the cognitive meaningfulness and/or truth-aptness of religious discourse. It is argued that this &ldquo;meta-level&rdquo; meaning agnosticism may actually be an attractive articulation of a certain kind of religious attitude. While pragmatists like William James have claimed that (epistemic) agnosticism practically collapses into atheism, meaning agnosticism at the meta-level can in fact be a pragmatist position focusing on our human condition and its limits. Additional issues, such as the relations between agnosticism and the theodicism vs. antitheodicism debate regarding the problem of evil and suffering, are also briefly examined.
  • Lajunen, Antti; Kivekäs, Klaus; Vepsäläinen, Jari; Tammi, Kari (Multidisciplinary Digital Publishing Institute, 2020)
    Different estimations have been presented for the amount of electric vehicles in the future. These estimations rarely take into account any realistic dynamics of the vehicle fleet. The objective of this paper is to analyze recently presented future scenarios about the passenger vehicle fleet estimations and create a foundation for the development of a fleet estimation model for passenger cars dedicated to the Finnish vehicle market conditions. The specific conditions of the Finnish light-duty vehicle fleet are taken into account as boundary conditions for the model development. The fleet model can be used for the estimation of emissions-optimal future vehicle fleets and the evaluation of the carbon dioxide emissions of transportation. The emission analysis was done for four different scenarios of the passenger vehicle fleet development in Finland. The results show that the high average age of the fleet and high number of older gasoline vehicles will slow down the reduction of carbon dioxide emissions during the next five to ten years even with a high adoption rate of electric vehicles. It can be concluded that lowering the average age, increasing biofuel mixing ratios, and increasing the amount of rechargeable electric vehicles are the most effective measures to reduce carbon dioxide emissions of the Finnish passenger vehicle fleet in the future.
  • Li, Yuhong; Su, Xiang; Ding, Aaron Yi; Lindgren, Anders; Liu, Xiaoli; Prehofer, Christian; Riekki, Jukka; Rahmani, Rahim; Tarkoma, Sasu; Hui, Pan (Multidisciplinary Digital Publishing Institute, 2020)
    The Internet of Things (IoT) connects smart devices to enable various intelligent services. The deployment of IoT encounters several challenges, such as difficulties in controlling and managing IoT applications and networks, problems in programming existing IoT devices, long service provisioning time, underused resources, as well as complexity, isolation and scalability, among others. One fundamental concern is that current IoT networks lack flexibility and intelligence. A network-wide flexible control and management are missing in IoT networks. In addition, huge numbers of devices and large amounts of data are involved in IoT, but none of them have been tuned for supporting network management and control. In this paper, we argue that Software-defined Networking (SDN) together with the data generated by IoT applications can enhance the control and management of IoT in terms of flexibility and intelligence. We present a review for the evolution of SDN and IoT and analyze the benefits and challenges brought by the integration of SDN and IoT with the help of IoT data. We discuss the perspectives of knowledge-driven SDN for IoT through a new IoT architecture and illustrate how to realize Industry IoT by using the architecture. We also highlight the challenges and future research works toward realizing IoT with the knowledge-driven SDN.
  • Rahman, Saifur; Alwadie, Abdullah S.; Irfan, Muhammed; Nawaz, Rabia; Raza, Mohsin; Javed, Ehtasham; Awais, Muhammad (Multidisciplinary Digital Publishing Institute, 2020)
    Gas sensors are critical components when adhering to health safety and environmental policies in various manufacturing industries, such as the petroleum and oil industry; scent and makeup production; food and beverage manufacturing; chemical engineering; pollution monitoring. In recent times, gas sensors have been introduced to medical diagnostics, bioprocesses, and plant disease diagnosis processes. There could be an adverse impact on human health due to the mixture of various gases (e.g., acetone (A), ethanol (E), propane (P)) that vent out from industrial areas. Therefore, it is important to accurately detect and differentiate such gases. Towards this goal, this paper presents a novel electronic nose (e-nose) detection method to classify various explosive gases. To detect explosive gases, metal oxide semiconductor (MOS) sensors are used as reliable tools to detect such volatile gases. The data received from MOS sensors are processed through a multivariate analysis technique to classify different categories of gases. Multivariate analysis was done using three variants&mdash;differential, relative, and fractional analyses&mdash;in principal components analysis (PCA). The MOS sensors also have three different designs: loading design, notch design, and Bi design. The proposed MOS sensor-based e-nose accurately detects and classifies three different gases, which indicates the reliability and practicality of the developed system. The developed system enables discrimination of these gases from the mixture. Based on the results from the proposed system, authorities can take preventive measures to deal with these gases to avoid their potential adverse impacts on employee health.
  • Mietzsch, Mario; McKenna, Robert; Väisänen, Elina; Yu, Jennifer C.; Ilyas, Maria; Hull, Joshua A.; Kurian, Justin; Smith, J. Kennon; Chipman, Paul; Lasanajak, Yi; Smith, David; Söderlund-Venermo, Maria; Agbandje-McKenna, Mavis (Multidisciplinary Digital Publishing Institute, 2020)
    Several members of the <i>Protoparvovirus</i> genus, capable of infecting humans, have been recently discovered, including cutavirus (CuV) and tusavirus (TuV). To begin the characterization of these viruses, we have used cryo-electron microscopy and image reconstruction to determine their capsid structures to ~2.9 &Aring; resolution, and glycan array and cell-based assays to identify glycans utilized for cellular entry. Structural comparisons show that the CuV and TuV capsids share common features with other parvoviruses, including an eight-stranded anti-parallel &beta;-barrel, depressions at the icosahedral 2-fold and surrounding the 5-fold axes, and a channel at the 5-fold axes. However, the viruses exhibit significant topological differences in their viral protein surface loops. These result in three separated 3-fold protrusions, similar to the bufaviruses also infecting humans, suggesting a host-driven structure evolution. The surface loops contain residues involved in receptor binding, cellular trafficking, and antigenic reactivity in other parvoviruses. In addition, terminal sialic acid was identified as the glycan potentially utilized by both CuV and TuV for cellular entry, with TuV showing additional recognition of poly-sialic acid and sialylated Lewis X (sLeXLeXLeX) motifs reported to be upregulated in neurotropic and cancer cells, respectively. These structures provide a platform for annotating the cellular interactions of these human pathogens.
  • Hellinen, Laura; Bahrpeyma, Sina; Rimpelä, Anna-Kaisa; Hagström, Marja; Reinisalo, Mika; Urtti, Arto (Multidisciplinary Digital Publishing Institute, 2020)
    Interactions between drugs and melanin pigment may have major impacts on pharmacokinetics. Therefore, melanin binding can modify the efficacy and toxicity of medications in ophthalmic and other disease of pigmented tissues, such as melanoma. As melanin is present in many pigmented tissues in the human body, investigation of pigment binding is relevant in drug discovery and development. Conventionally, melanin binding assays have been performed using an equilibrium binding study followed by chemical analytics, such as LC/MS. This approach is laborious, relatively slow, and limited to facilities with high performance quantitation instrumentation. We present here a screening of melanin binding with label-free microscale thermophoresis (MST) that utilizes the natural autofluorescence of melanin. We determined equilibrium dissociation constants (K<sub>d</sub>) of 11 model compounds with melanin nanoparticles. MST categorized the compounds into extreme (chloroquine, penicillin G), high (papaverine, levofloxacin, terazosin), intermediate (timolol, nadolol, quinidine, propranolol), and low melanin binders (atropine, methotrexate, diclofenac) and displayed good correlation with binding parameter values obtained with the conventional binding study and LC/MS analytics. Further, correlation was seen between predicted melanin binding in human retinal pigment epithelium and choroid (RPE-choroid) and K<sub>d</sub> values obtained with MST. This method represents a useful and fast approach for classification of compounds regarding melanin binding. Thus, the method can be utilized in various fields, including drug discovery, pharmacokinetics, and toxicology.
  • Mehmood, Furrukh; Abdullah,; Ubaid, Zartasha; Bao, Yiming; Poczai, Peter; Mirza, Bushra (Multidisciplinary Digital Publishing Institute, 2020)
    Within the family Solanaceae, <i>Withania</i> is a small genus belonging to the Solanoideae subfamily. Here, we report the de novo assembled chloroplast genome sequences of <i>W. coagulans, W. adpressa</i>, and<i> W. riebeckii. </i>The length of these genomes ranged from 154,162 to 154,364 base pairs (bp). These genomes contained a pair of inverted repeats (IRa and IRb) ranging from 25,029 to 25,071 bp that were separated by a large single-copy (LSC) region of 85,635&ndash;85,765 bp and a small single-copy (SSC) region of 18,457&ndash;18,469 bp. We analyzed the structural organization, gene content and order, guanine-cytosine content, codon usage, RNA-editing sites, microsatellites, oligonucleotide and tandem repeats, and substitutions of <i>Withania</i> plastomes, which revealed high similarities among the species. Comparative analysis among the <i>Withania</i> species also highlighted 10 divergent hotspots that could potentially be used for molecular marker development, phylogenetic analysis, and species identification. Furthermore, our analyses showed that even three mutational hotspots (<i>rps</i>4-<i>trn</i>T, <i>trn</i>M-<i>atp</i>E, and <i>rps</i>15) were sufficient to discriminate the <i>Withania</i> species included in current study.
  • Abdullah,; Henriquez, Claudia L.; Mehmood, Furrukh; Shahzadi, Iram; Ali, Zain; Waheed, Mohammad Tahir; Croat, Thomas B.; Poczai, Peter; Ahmed, Ibrar (Multidisciplinary Digital Publishing Institute, 2020)
    The chloroplast genome provides insight into the evolution of plant species. We de novo assembled and annotated chloroplast genomes of four genera representing three subfamilies of Araceae: <i>Lasia</i> <i>spinosa</i> (Lasioideae), <i>Stylochaeton bogneri</i>, <i>Zamioculcas zamiifolia</i> (Zamioculcadoideae), and <i>Orontium</i> <i>aquaticum</i> (Orontioideae), and performed comparative genomics using these chloroplast genomes. The sizes of the chloroplast genomes ranged from 163,770 bp to 169,982 bp. These genomes comprise 113 unique genes, including 79 protein-coding, 4 rRNA, and 30 tRNA genes. Among these genes, 17&ndash;18 genes are duplicated in the inverted repeat (IR) regions, comprising 6&ndash;7 protein-coding (including trans-splicing gene <i>rps</i>12), 4 rRNA, and 7 tRNA genes. The total number of genes ranged between 130 and 131. The <i>inf</i>A gene was found to be a pseudogene in all four genomes reported here. These genomes exhibited high similarities in codon usage, amino acid frequency, RNA editing sites, and microsatellites. The oligonucleotide repeats and junctions JSB (IRb/SSC) and JSA (SSC/IRa) were highly variable among the genomes. The patterns of IR contraction and expansion were shown to be homoplasious, and therefore unsuitable for phylogenetic analyses. Signatures of positive selection were seen in three genes in <i>S. bogneri,</i> including <i>ycf</i>2, <i>clp</i>P, and <i>rpl</i>36. This study is a valuable addition to the evolutionary history of chloroplast genome structure in Araceae.
  • Ahlgren, Tommy; Heinola, Kalle (Multidisciplinary Digital Publishing Institute, 2020)
    The application of mean-field rate theory equations have proven to be a versatile method in simulating defect dynamics and temporal changes in the microstructure of materials. The reliability and usefulness of the method, however, depends critically on the defect interaction parameters used. In this study, we show that the main interaction parameter, the sink strength, intrinsically depends on the detrapping, or the dissociation process itself. We present a theory on how to determine the appropriate sink strengths. The correct sink strength required for a detrapping defect, is considerably larger than the values commonly used, and thus should not be neglected.
  • Jiang, Zhong-Qing; Wang, Jing; Stoddard, Frederick; Salovaara, Hannu; Sontag-Strohm, Tuula (Multidisciplinary Digital Publishing Institute, 2020)
    Faba bean protein has good functionalities, but it is little used in the food industry. This study identified a challenge from unfavourable starch gelation when utilizing faba bean for producing protein-based emulsion gel foods, and developed processing methods to overcome that. Two types of protein-based emulsion gel foods, namely yogurt and tofu analogue products, were prepared. The processing methods in this study involved steps of thermal pre-treatment of the beans, dehulling, milling, adding plant oil, homogenization, prevention of starch gelation, and inducing protein gelation. Two methods for preventing starch gelation were studied, namely starch removal and hydrolysis. The gel texture, water-holding capacity, and structural properties of the gel products were evaluated. Both starch-gelation prevention methods produced yogurt and tofu analogue products having typical emulsion gel properties. Hydrolysis of starch was favourable for producing the yogurt analogue, because the hydrolysate compounds improved the gel strength and viscosity. Moreover, it utilized the whole flour, meaning all the nutrients from the cotyledon were used and no side-stream was created. In contrast, starch removal was slightly better than hydrolysis for producing the tofu analogue, because the hydrolysate lowered the gel strength and water-holding capacity of the products. It is both possible and ecologically sustainable to utilize whole faba bean flour for making emulsion gel products.
  • Leskinen, Katarzyna; Pajunen, Maria I.; Vilanova, Miguel Vincente Gomez-Raya; Kiljunen, Saija; Nelson, Andrew; Smith, Darren; Skurnik, Mikael (Multidisciplinary Digital Publishing Institute, 2020)
    YerA41 is a <i>Myoviridae</i> bacteriophage that was originally isolated due its ability to infect <i>Yersinia ruckeri</i> bacteria, the causative agent of enteric redmouth disease of salmonid fish. Several attempts to determine its genomic DNA sequence using traditional and next generation sequencing technologies failed, indicating that the phage genome is modified in such a way that it is an unsuitable template for PCR amplification and for conventional sequencing. To determine the YerA41 genome sequence, we performed RNA-sequencing from phage-infected <i>Y. ruckeri</i> cells at different time points post-infection. The host-genome specific reads were subtracted and de novo assembly was performed on the remaining unaligned reads. This resulted in nine phage-specific scaffolds with a total length of 143 kb that shared only low level and scattered identity to known sequences deposited in DNA databases. Annotation of the sequences revealed 201 predicted genes, most of which found no homologs in the databases. Proteome studies identified altogether 63 phage particle-associated proteins. The RNA-sequencing data were used to characterize the transcriptional control of YerA41 and to investigate its impact on the bacterial gene expression. Overall, our results indicate that RNA-sequencing can be successfully used to obtain the genomic sequence of non-sequencable phages, providing simultaneous information about the phage–host interactions during the process of infection.
  • Jaakkonen, Anniina; Volkmann, Gerrit; Iwaï, Hideo (Multidisciplinary Digital Publishing Institute, 2020)
    Monoclonal antibodies, engineered antibodies, and antibody fragments have become important biological therapeutic platforms. The IgG format with bivalent binding sites has a modular structure with different biological roles, i.e., effector and binding functions, in different domains. We demonstrated the reconstruction of an IgG-like domain structure in vitro by protein ligation using protein <i>trans</i>-splicing. We produced various binding domains to replace the binding domain of IgG from <i>Escherichia coli</i> and the Fc domain of human IgG from <i>Brevibacillus choshinensis</i> as split-intein fusions. We showed that in vitro protein ligation could produce various Fc-fusions at the N-terminus in vitro from the independently produced domains from different organisms. We thus propose an off-the-shelf approach for the combinatorial production of Fc fusions in vitro with several distinct binding domains, particularly from naturally occurring binding domains. Antiviral lectins from algae are known to inhibit virus entry of HIV and SARS coronavirus. We demonstrated that a lectin could be fused with the Fc-domain in vitro by protein ligation, producing an IgG-like molecule as a &ldquo;lectibody&rdquo;. Such an Fc-fusion could be produced in vitro by this approach, which could be an attractive method for developing potential therapeutic agents against rapidly emerging infectious diseases like SARS coronavirus without any genetic fusion and expression optimization.

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