Sustainable production of eicosapentaenoic acid-rich oil from microalgae: Towards an algal biorefinery

Utilization of sustainable natural resources such as microalgae has been considered for the production of biofuels, aquaculture feed, high-value bioactives such as omega-3 fatty acids, carotenoids, etc. Eicosapentaenoic acid (EPA) is an omega-3 fatty acid present in fish oil, which is of physiological importance to both humans and fish. Marine microalgae are sustainable sources of lipid rich in EPA, and different species have been explored for the production of EPA as a single product. There has been a rising interest in the concept of a multi-product biorefinery, focusing on the maximum valorization of the algal biomass. Targeting one or more value-added compounds in a biorefinery scenario can improve the commercial viability of low-value products such as triglycerides for biofuel. This approach has been viewed by technologists and experts as a sustainable and economically feasible possibility for the large-scale production of microalgae for its potential applications in biodiesel and jet fuel production, nutraceuticals, animal and aquaculture feeds, etc. In this review paper, we describe the recent developments in the production of high-value EPA-rich oil from microalgae, emphasizing the upstream and downstream bioprocess techniques, and the advantages of considering an EPA-rich oil-based biorefinery.     

Differential effects of anti-cancer and anti-hepatitis drugs on liver cystatin

The drug–protein interaction has been the subject of increasing interest over the decades. In the present communication, the interaction of liver cystatin with anti-cancer (adriamycin) and anti-hepatitis (adevofir dipivoxil) drugs was studied by thiol-protease inhibitory assay, UV absorption, fluorescence spectroscopy and circular dichroism (CD). A static type of quenching was observed between the protein and the drug molecules. Binding constant (Ka) of adriamycin to liver cystatin (LC) was found to be 1.08 × 10⁶ M⁻¹. Moreover, binding site number was found to be 2. Importantly, cystatin loses its activity in the presence of adriamycin. However, intrinsic fluorescence studies in the presence of adevofir dipivoxil showed enhancement in the fluorescence intensity suggesting that binding of adevofir to LC caused unfolding of the protein. The unfolding of the test protein was also accompanied by significant loss of inhibitory activity. CD spectroscopy result showed, both adriamycin and adevofir dipivoxil caused perturbation in the secondary structure of liver cystatin. The possible implications of these results will help in combating drug induced off target effects.Abbreviations: LC, liver cystatin; ADR, adriamycin; CD, circular dichroism; Ka, binding constant; HBV, human hepatitis B virus     

Apoptosis induced by crocidolite asbestos in human lung epithelial cells involves inactivation of Akt and MAPK pathways

Exposure of human lung epithelial (A549) cells to asbestos fibers causes apoptosis, which is largely attributed to release of iron and generation of reactive oxygen species (ROS) within the cells. To mimic the highly oxidative environment generated by asbestos exposure in the absence of the actual fibers, we used two chemicals; buthione sulfoximine (BSO), an inhibitor of glutathione (GSH) synthesis and ferric ammonium citrate (FAC), a source of iron. Here, we report that exposure of A549 cells to crocidolite asbestos led to a significant time-dependent inactivation of signaling proteins, i.e. Akt and all mitogen-activated protein kinases (MAPKs) (p38, ERK1/2 and SAPK/JNK), and subsequently to apoptosis. Unlike crocidolite treatment, the use of BSO and FAC, independently or combined, did not change the phosphorylation status of proteins, nor did it induce apoptosis. Taken together, our results presented herein point to the possibility that crocidolite-induced apoptosis of human lung epithelial cells is not a mere consequence of generation of oxidants but also requires inactivation of major cell growth and differentiation pathways. A. Baldys, P. Pande contributed equally to this publication.     

A systematic analysis of eluted fraction of plasma post immunoaffinity depletion: implications in biomarker discovery

Plasma is the most easily accessible source for biomarker discovery in clinical proteomics. However, identifying potential biomarkers from plasma is a challenge given the large dynamic range of proteins. The potential biomarkers in plasma are generally present at very low abundance levels and hence identification of these low abundance proteins necessitates the depletion of highly abundant proteins. Sample pre-fractionation using immuno-depletion of high abundance proteins using multi-affinity removal system (MARS) has been a popular method to deplete multiple high abundance proteins. However, depletion of these abundant proteins can result in concomitant removal of low abundant proteins. Although there are some reports suggesting the removal of non-targeted proteins, the predominant view is that number of such proteins is small. In this study, we identified proteins that are removed along with the targeted high abundant proteins. Three plasma samples were depleted using each of the three MARS (Hu-6, Hu-14 and Proteoprep 20) cartridges. The affinity bound fractions were subjected to gelC-MS using an LTQ-Orbitrap instrument. Using four database search algorithms including MassWiz (developed in house), we selected the peptides identified at <1% FDR. Peptides identified by at least two algorithms were selected for protein identification. After this rigorous bioinformatics analysis, we identified 101 proteins with high confidence. Thus, we believe that for biomarker discovery and proper quantitation of proteins, it might be better to study both bound and depleted fractions from any MARS depleted plasma sample.     

Unraveling the Web of Viroinformatics: Computational Tools and Databases in Virus Research

The beginning of the second century of research in the field of virology (the first virus was discovered in 1898) was marked by its amalgamation with bioinformatics, resulting in the birth of a new domain—viroinformatics. The availability of more than 100 Web servers and databases embracing all or specific viruses (for example, dengue virus, influenza virus, hepatitis virus, human immunodeficiency virus [HIV], hemorrhagic fever virus [HFV], human papillomavirus [HPV], West Nile virus, etc.) as well as distinct applications (comparative/diversity analysis, viral recombination, small interfering RNA [siRNA]/short hairpin RNA [shRNA]/microRNA [miRNA] studies, RNA folding, protein-protein interaction, structural analysis, and phylotyping and genotyping) will definitely aid the development of effective drugs and vaccines. However, information about their access and utility is not available at any single source or on any single platform. Therefore, a compendium of various computational tools and resources dedicated specifically to virology is presented in this article.     

Excision of HIV-1 proviral DNA by recombinant cell permeable tre-recombinase

Over the previous years, comprehensive studies on antiretroviral drugs resulted in the successful introduction of highly active antiretroviral therapy (HAART) into clinical practice for treatment of HIV/AIDS. However, there is still need for new therapeutic approaches, since HAART cannot eradicate HIV-1 from the infected organism and, unfortunately, can be associated with long-term toxicity and the development of drug resistance. In contrast, novel gene therapy strategies may have the potential to reverse the infection by eradicating HIV-1. For example, expression of long terminal repeat (LTR)-specific recombinase (Tre-recombinase) has been shown to result in chromosomal excision of proviral DNA and, in consequence, in the eradication of HIV-1 from infected cell cultures. However, the delivery of Tre-recombinase currently depends on the genetic manipulation of target cells, a process that is complicating such therapeutic approaches and, thus, might be undesirable in a clinical setting. In this report we demonstrate that E.coli expressed Tre-recombinases, tagged either with the protein transduction domain (PTD) from the HIV-1 Tat trans-activator or the translocation motif (TLM) of the Hepatitis B virus PreS2 protein, were able to translocate efficiently into cells and showed significant recombination activity on HIV-1 LTR sequences. Tre activity was observed using episomal and stable integrated reporter constructs in transfected HeLa cells. Furthermore, the TLM-tagged enzyme was able to excise the full-length proviral DNA from chromosomal integration sites of HIV-1-infected HeLa and CEM-SS cells. The presented data confirm Tre-recombinase activity on integrated HIV-1 and provide the basis for the non-genetic transient application of engineered recombinases, which may be a valuable component of future HIV eradication strategies.     

The l2 minor capsid protein of human papillomavirus type 16 interacts with a network of nuclear import receptors

The L2 minor capsid proteins enter the nucleus twice during viral infection: in the initial phase after virion disassembly and in the productive phase when, together with the L1 major capsid proteins, they assemble the replicated viral DNA into virions. In this study we investigated the interactions between the L2 protein of high-risk human papillomavirus type 16 (HPV16) and nuclear import receptors. We discovered that HPV16 L2 interacts directly with both Kapbeta(2) and Kapbeta(3). Moreover, binding of Ran-GTP to either Kapbeta(2) or Kapbeta(3) inhibits its interaction with L2, suggesting that the Kapbeta/L2 complex is import competent. In addition, we found that L2 forms a complex with the Kapalpha(2)beta(1) heterodimer via interaction with the Kapalpha(2) adapter. In agreement with the binding data, nuclear import of L2 in digitonin-permeabilized cells could be mediated by either Kapalpha(2)beta(1) heterodimers, Kapbeta(2), or Kapbeta(3). Mapping studies revealed that HPV16 L2 contains two nuclear localization signals (NLSs), in the N terminus (nNLS) and C terminus (cNLS), that could mediate its nuclear import. Together the data suggest that HPV16 L2 interacts via its NLSs with a network of karyopherins and can enter the nucleus via several import pathways mediated by Kapalpha(2)beta(1) heterodimers, Kapbeta(2), and Kapbeta(3).     

Perceptions and preferences on denture marking in an Indian sample

doi: 10.1111/j.1741‐2358.2011.00499.x Perceptions and preferences on denture marking in an Indian sample Aim: Denture marking is useful in institutional settings and post‐mortem identification. Numerous markers have been developed, and their advantages and limitations assessed previously; however, patient perception to denture marking is paramount. We evaluated this in an Indian sample and also gauged their preference for different markers. Materials and methods: One‐hundred and one edentulous patients seeking prosthodontic treatment in our institution were shown four denture markers (stainless steel matrix band, paper strip with name inscribed on it, patient photograph and optically readable laminated bar code) and asked whether they wanted similar markers in their dentures; patients were also asked to rank the markers based on preference and indicate their satisfaction with it. Results: Approximately two‐thirds of patients (65/101) were uninterested in getting their dentures marked; among the 36 who agreed, 10 preferred the stainless steel band followed by photographs (9), paper strip (6) and bar code (2); nine gave multiple responses and were excluded from analyses. Sixteen patients expressed dissatisfaction with the photographic marker and bar code, while this number reduced for the stainless steel band (13) and paper strip (10). Conclusions: The results are in contrast to European studies wherein the majority of patients agreed to denture marking, indicating patient background (e.g. education level) may affect perception to denture marking; amongst those who agreed to marking, most preferred, or were satisfied with, the stainless steel and paper strip markers over photographic marker and a complex method such as bar‐coding, implying that simple methods harbouring basic patient information may suffice in the Indian context.