Anwar Sunna

Cell-Free Biocatalysis for the Production of Platform Chemicals

Publisher: Frontiers Media SA

Date: 27-08-2020

DOI: 10.3389/FENRG.2020.00193

Prospecting for novel lipase genes using PCR a aThe GenBank accession number for the seque

Publisher: Microbiology Society

Date: 08-2002

DOI: 10.1099/00221287-148-8-2283

Abstract: A PCR method suitable for the isolation of lipase genes directly from environmental DNA is described. The problems associated with the low levels of similarity between lipase genes were overcome by extensive analysis of conserved regions and careful primer design. Using this method, a lipase gene (oli-lipase) was isolated directly from environmental DNA. This lipase showed less than 20% similarity with other known lipases at the amino acid level. The study also revealed that distantly related members of the alpha/beta hydrolase superfamily share similar conserved motifs with the lipases, thus making these genes targets for gene prospecting by PCR.

Construction of a cell-free synthetic pathway for the production of lactic acid from spent coffee grounds

Publisher: Elsevier BV

Date: 10-2018

DOI: 10.1016/J.NBT.2018.05.872

Biofunctionalization of silica-coated magnetic particles mediated by a peptide

Publisher: Springer Science and Business Media LLC

Date: 18-07-2014

DOI: 10.1007/S11051-014-2543-7

A comprehensive assessment of the biosynthetic pathways of ascorbate, α-tocopherol and free amino acids in Euglena gracilis var. saccharophila

Publisher: Elsevier BV

Date: 11-2017

DOI: 10.1016/J.ALGAL.2017.08.029

Probing the Role of the Chloroplasts in Heavy Metal Tolerance and Accumulation in Euglena gracilis

Publisher: MDPI AG

Date: 14-01-2020

DOI: 10.3390/MICROORGANISMS8010115

Abstract: The E. gracilis Zm-strain lacking chloroplasts, characterized in this study, was compared with the earlier assessed wild type Z-strain to explore the role of chloroplasts in heavy metal accumulation and tolerance. Comparison of the minimum inhibitory concentration (MIC) values indicated that both strains tolerated similar concentrations of mercury (Hg) and lead (Pb), but cadmium (Cd) tolerance of the Z-strain was twice that of the Zm-strain. The ability of the Zm-strain to accumulate Hg was higher compared to the Z-strain, indicating the existence of a Hg transportation and accumulation mechanism not depending on the presence of chloroplasts. Transmission electron microscopy (TEM) showed maximum accumulation of Hg in the cytosol of the Zm-strain and highest accumulation of Cd in the chloroplasts of the Z-strain indicating a difference in the ability of the two strains to deposit heavy metals in the cell. The highly abundant heavy metal transporter MTP2 in the Z-strain may have a role in Cd transportation to the chloroplasts. A multidrug resistance-associated protein highly increased in abundance in the Zm-strain could be a potential Hg transporter to either cytosol or mitochondria. Overall, the chloroplasts appear to have major role in the tolerance and accumulation of Cd in E. gracilis.

Comparative proteomics investigation of central carbon metabolism in Euglena gracilis grown under predominantly phototrophic, mixotrophic and heterotrophic cultivations

Publisher: Elsevier BV

Date: 11-2019

DOI: 10.1016/J.ALGAL.2019.101638

Bioengineering Strategies for Protein-Based Nanoparticles

Publisher: MDPI AG

Date: 23-07-2018

DOI: 10.3390/GENES9070370

Abstract: In recent years, the practical application of protein-based nanoparticles (PNPs) has expanded rapidly into areas like drug delivery, vaccine development, and biocatalysis. PNPs possess unique features that make them attractive as potential platforms for a variety of nanobiotechnological applications. They self-assemble from multiple protein subunits into hollow monodisperse structures they are highly stable, biocompatible, and biodegradable and their external components and encapsulation properties can be readily manipulated by chemical or genetic strategies. Moreover, their complex and perfect symmetry have motivated researchers to mimic their properties in order to create de novo protein assemblies. This review focuses on recent advances in the bioengineering and bioconjugation of PNPs and the implementation of synthetic biology concepts to exploit and enhance PNP’s intrinsic properties and to impart them with novel functionalities.

Solid-Binding Peptides: Immobilisation Strategies for Extremophile Biocatalysis in Biotechnology

Publisher: Springer International Publishing

Date: 2016

DOI: 10.1007/978-3-319-13521-2_23

Effect of Combination l-Citrulline and Metformin Treatment on Motor Function in Patients With Duchenne Muscular Dystrophy

Publisher: American Medical Association (AMA)

Date: 30-10-2019

DOI: 10.1001/JAMANETWORKOPEN.2019.14171

Linker-protein G mediated functionalization of polystyrene-encapsulated upconversion nanoparticles for rapid gene assay using convective PCR.

Publisher: Springer Science and Business Media LLC

Date: 11-05-2019

DOI: 10.1007/S00604-019-3466-X

Abstract: The authors report on a simplified approach to encapsulate upconversion nanoparticles (UCNPs) in polystyrene spheres by mini-emulsion polymerisation. The resulting particles (PS-UCNP) are hydrophilic, stable and suitable for biomolecular recognition and biosensing applications. Also, a strategy was developed for bioconjugation of antibodies onto the surface of the PS-UCNPs by using the bifunctional fusion protein linker-protein G (LPG). LPG mediates the functionalisation of PS-UCNPs with antibodies against digoxigenin allowing for specific labelling of convective PCR (cPCR) licons. Lambda DNA was lified using cPCR on a heat block for 30 min using the digoxigenin labelled forward and biotin labelled reverse primers. The antibody functionalised PS-UCNPs bind to the digoxigenin end of the cPCR licons. Finally, the streptavidin labelled magnetic beads were used to selectively capture the PS-UCNP-labelled cPCR licons and the upconversion signal was detected at 537 nm under 980 nm excitation. This sandwich approach enables direct recognition of the target lambda DNA with a detection limit of 10

Elucidating the Binding Mechanism of a Novel Silica-Binding Peptide

Publisher: MDPI AG

Date: 18-12-2019

DOI: 10.3390/BIOM10010004

Abstract: Linker-protein G (LPG) is a bifunctional fusion protein composed of a solid-binding peptide (SBP, referred as the “linker”) with high affinity to silica-based compounds and a Streptococcus protein G (PG), which binds antibodies. The binding mechanisms of LPG to silica-based materials was studied using different biophysical techniques and compared to that of PG without the linker. LPG displayed high binding affinity to a silica surface (KD = 34.77 ± 11.8 nM), with a vertical orientation, in comparison to parent PG, which exhibited no measurable binding affinity. Incorporation of the linker in the fusion protein, LPG, had no effect on the antibody-binding function of PG, which retained its secondary structure and displayed no alteration of its chemical stability. The LPG system provided a milder, easier, and faster affinity-driven immobilization of antibodies to inorganic surfaces when compared to traditional chemical coupling techniques.

Solid-binding peptides: Smart tools for nanobiotechnology

Publisher: Elsevier BV

Date: 05-2015

DOI: 10.1016/J.TIBTECH.2015.02.005

Abstract: Over the past decade, solid-binding peptides (SBPs) have been used increasingly as molecular building blocks in nanobiotechnology. These peptides show selectivity and bind with high affinity to the surfaces of a erse range of solid materials including metals, metal oxides, metal compounds, magnetic materials, semiconductors, carbon materials, polymers, and minerals. They can direct the assembly and functionalisation of materials, and have the ability to mediate the synthesis and construction of nanoparticles and complex nanostructures. As the availability of newly synthesised nanomaterials expands rapidly, so too do the potential applications for SBPs.

A novel universal detection agent for time-gated luminescence bioimaging

Publisher: Springer Science and Business Media LLC

Date: 10-06-2016

DOI: 10.1038/SREP27564

Abstract: Luminescent lanthanide chelates have been used to label antibodies in time-gated luminescence (TGL) bioimaging. However, it is a challenging task to label directly an antibody with lanthanide-binding ligands and achieve control of the target ligand rotein ratios whilst ensuring that affinity and avidity of the antibody remain uncompromised. We report the development of a new indirect detection reagent to label antibodies with detectable luminescence that circumvents this problem by labelling available lysine residues in the linker portion of the recombinant fusion protein Linker-Protein G (LPG). Succinimide-activated lanthanide chelating ligands were attached to lysine residues in LPG and Protein G (without Linker) and the resulting Luminescence-Activating (LA-) conjugates were compared for total incorporation and conjugation efficiency. A higher and more efficient incorporation of ligands at three different molar ratios was observed for LPG and this effect was attributed to the presence of eight readily available lysine residues in the linker region of LPG. These Luminescence-Activating (LA-) complexes were subsequently shown to impart luminescence (upon formation of europium(III) complexes) to cell-specific antibodies within seconds and without the need for any complicated bioconjugation procedures. The potential of this technology was demonstrated by direct labelling of Giardia cysts and Cryptosporidium oocysts in TGL bioimaging.

Bioengineering a light-responsive encapsulin nanoreactor

Publisher: American Chemical Society (ACS)

Date: 15-02-2021

DOI: 10.1021/ACSAMI.0C21141

Characterisation of the first archaeal mannonate dehydratase from Thermoplasma acidophilum and its potential role in the catabolism of D-Mannose

Publisher: MDPI AG

Date: 03-03-2019

DOI: 10.3390/CATAL9030234

Abstract: Mannonate dehydratases catalyse the dehydration reaction from mannonate to 2-keto-3-deoxygluconate as part of the hexuronic acid metabolism in bacteria. Bacterial mannonate dehydratases present in this gene cluster usually belong to the xylose isomerase-like superfamily, which have been the focus of structural, biochemical and physiological studies. Mannonate dehydratases from archaea have not been studied in detail. Here, we identified and characterised the first archaeal mannonate dehydratase (TaManD) from the thermoacidophilic archaeon Thermoplasma acidophilum. The recombinant TaManD enzyme was optimally active at 65 °C and showed high specificity towards D-mannonate and its lactone, D-mannono-1,4-lactone. The gene encoding for TaManD is located adjacent to a previously studied mannose-specific aldohexose dehydrogenase (AldT) in the genome of T. acidophilum. Using nuclear magnetic resonance (NMR) spectroscopy, we showed that the mannose-specific AldT produces the substrates for TaManD, demonstrating the possibility for an oxidative metabolism of mannose in T. acidophilum. Among previously studied mannonate dehydratases, TaManD showed closest homology to enzymes belonging to the xylose isomerase-like superfamily. Genetic analysis revealed that closely related mannonate dehydratases among archaea are not located in a hexuronate gene cluster like in bacteria, but next to putative aldohexose dehydrogenases, implying a different physiological role of mannonate dehydratases in those archaeal species.

Smartphone detection of antibiotic resistance using convective PCR and a lateral flow assay

Publisher: Elsevier BV

Date: 11-2019

DOI: 10.1016/J.SNB.2019.126849

Adapted method for rapid detection and quantification of pathogen Campylobacter jejuni from environmental water samples

Publisher: Oxford University Press (OUP)

Date: 27-05-2023

DOI: 10.1093/FEMSEC/FIAD058

Abstract: Building on a previously developed workflow for rapid and sensitive pathogen detection by qPCR, this work has established a s le treatment strategy that produces consistent quantification efficiencies (QEs) for C ylobacter jejuni against a complex and highly variable s le matrix from a suburban river. The in idual treatments most effective at minimizing the inhibitory effects of the s le matrix were pH buffering with HEPES (50 mM, pH 5.7) and addition of the surfactant Tween 20 (2% v/v). Unexpectedly, s le acidification (pH 4–5) resulting from the use of aged Tween 20 that had undergone partial hydrolysis, appeared to play a key role in enhancing QE. This effect could be replicated by direct pH adjustment with dilute hydrochloric acid and may be linked to the solubilization and removal of inhibitory particles at an acidic pH. While the effectiveness of each in idual treatment method varied, a combined treatment of either HEPES buffer + Tween 20, or direct pH adjustment + Tween 20, consistently produced QEs of 60%–70% and up to 100%, respectively, over a s ling period of one year. The consistency and scalability of this workflow make it a suitable alternative to culture-based ISO methods for detecting C ylobacter spp.

Peptides and peptide-based biomaterials and their biomedical applications

Publisher: Springer International Publishing

Date: 2017

DOI: 10.1007/978-3-319-66095-0

Effect of Trichoderma reesei proteinases on the affinity of an inorganic-binding peptide

Publisher: Springer Science and Business Media LLC

Date: 27-06-2014

DOI: 10.1007/S12010-014-1027-7

Abstract: An inorganic-binding peptide sequence with high affinity to silica-containing materials was fused to a glycoside hydrolase GH26 mannanase, ManA, from the extremely thermophilic bacterium Dictyoglomus thermophilum. The resulting recombinant enzyme produced in Escherichia coli, ManA-Linker, displayed high binding affinity towards synthetic zeolite while retaining its catalytic activity at 80 °C. ManA-Linker was able to bind to the zeolite at different pH levels, indicating a true pH-independent binding. However, complete degradation of the peptide linker was observed when the recombinant ManA-Linker was exposed to the supernatant from the filamentous fungus Trichoderma reesei. This degradation was caused by extracellular proteinases produced by T. reesei during its growth phase. Several derivatives of ManA-Linker were designed and expressed in E. coli. All the derivatives carrying a single sequence of the linker were still susceptible to T. reesei proteinase degradation. Complete substitution of the linker sequence by (GGGGS)16 resulted in a proteinase-resistant ManA derivative, ManA-Linker-(GGGGS)16, which was able to bind to zeolite in a pH-dependent manner.

The Effect of Oligomerization on A Solid-Binding Peptide Binding to Silica-Based Materials

Publisher: MDPI AG

Date: 30-05-2020

DOI: 10.3390/NANO10061070

Abstract: The bifunctional linker-protein G (LPG) fusion protein comprises a peptide (linker) sequence and a truncated form of Streptococcus strain G148 protein G (protein G). The linker represents a multimeric solid-binding peptide (SBP) comprising 4 × 21-amino acid sequence repeats that display high binding affinity towards silica-based materials. In this study, several truncated derivatives were investigated to determine the effect of the SBP oligomerization on the silica binding function of LPG (for the sake of clarity, LPG will be referred from here on as 4 × LPG). Various biophysical characterization techniques were used to quantify and compare the truncated derivatives against 4 × LPG and protein G without linker (PG). The derivative containing two sequence repeats (2 × LPG) showed minimal binding to silica, while the truncated derivative with only a single sequence (1 × LPG) displayed no binding. The derivative containing three sequence repeats (3 × LPG) was able to bind to silica with a binding affinity of KD = 53.23 ± 4.5 nM, which is 1.5 times lower than that obtained for 4 × LPG under similar experimental conditions. Circular dichroism (CD) spectroscopy and fluorescence spectroscopy studies indicated that the SBP degree of oligomerization has only a small effect on the secondary structure (the linker unravels the beginning of the protein G sequence) and chemical stability of the parent protein G. However, based on quartz crystal microbalance with dissipation monitoring (QCM-D), oligomerization is an important parameter for a strong and stable binding to silica. The replacement of three sequence repeats by a (GGGGS)12 glycine-rich spacer indicated that the overall length rather than the SBP oligomerization mediated the effective binding to silica.

Versatile Platform for Nanoparticle Surface Bioengineering Based on SiO

Publisher: American Chemical Society (ACS)

Date: 27-04-2018

DOI: 10.1021/ACSAMI.8B01627

Abstract: Nanoparticle surface engineering can change its chemical identity to enable surface coupling with functional biomolecules. However, common surface coupling methods such as physical adsorption or chemical conjugation often suffer from the low coupling yield, poorly controllable orientation of biomolecules, and steric hindrance during target binding. These issues limit the application scope of nanostructures for theranostics and personalized medicine. To address these shortfalls, we developed a rapid and versatile method of nanoparticle biomodification. The method is based on a SiO

A novel framework for the cell-free enzymatic production of glucaric acid

Publisher: Elsevier BV

Date: 2020

DOI: 10.1016/J.YMBEN.2019.11.003

Abstract: Glucaric acid (GlucA) is a valuable glucose-derived chemical with promising applications as a biodegradable and biocompatible chemical in the manufacturing of plastics, detergents and drugs. Recently, there has been a significant focus on producing GlucA microbially (in vivo) from renewable materials such as glucose, sucrose and myo-inositol. However, these in vivo GlucA production processes generally lack efficiency due to toxicity problems, metabolite competition and suboptimal enzyme ratios. Synthetic biology and accompanying cell-free biocatalysis have been proposed as a viable approach to overcome many of these limitations. However, cell-free biocatalysis faces its own limitations for industrial applications due to high enzyme costs and cofactor consumption. We have constructed a cell-free GlucA pathway and demonstrated a novel framework to overcome limitations of cell-free biocatalysis by i) the combination of both thermostable and mesophilic enzymes, ii) incorporation of a cofactor regeneration system and iii) immobilisation and recycling of the pathway enzymes. The cell-free production of GlucA was achieved from glucose-1-phosphate with a titre of 14.1 ± 0.9 mM (3.0 ± 0.2 g l

Mixed-mode liquid chromatography for the rapid analysis of biocatalytic glucaric acid reaction pathways.

Publisher: Elsevier BV

Date: 08-2019

DOI: 10.1016/J.ACA.2019.03.023

Abstract: Glucaric acid (GlucA) has been identified as one of the top 10 potential bio-based chemicals for replacement of oil-based chemicals. Several synthetic enzyme pathways have been engineered in bacteria and yeast to produce GlucA from glucose and myo-inositol. However, the yields and titres achieved with these systems remain too low for the requirements of a bio-based GlucA industry. A major limitation for the optimisation of GlucA production via synthetic enzymatic pathways are the laborious analytical procedures required to detect the final product (GlucA) and pathway intermediates. We have developed a novel method for the simple and simultaneous analysis of GlucA and pathway intermediates to address this limitation using mixed mode (MM) HILIC and weak anion exchange chromatography (WAX), referred to as MM HILIC/WAX, coupled with RID. Isocratic mobile phase conditions and the s le solvent were optimised for the separation of GlucA, glucose-1-phosphate (G1P), glucose-6-phosphate (G6P), inositol-1-phosphate (I1P), myo-inositol and glucuronic acid (GA). The method showed good repeatability, precision and excellent accuracy with detection and quantitation limits (LOD and LOQ) of 1.5-2 and 577 mM, respectively. The method developed was used for monitoring the enzymatic synthesis of the final step in the GlucA pathway, and showed that GlucA was produced from GA with near 100% conversion and a titre of 9.2 g L

Universal Enzyme-Based Field Workflow for Rapid and Sensitive Quantification of Water Pathogens

Publisher: MDPI AG

Date: 16-11-2021

DOI: 10.3390/MICROORGANISMS9112367

Abstract: A universal filtration and enzyme-based workflow has been established to allow for the rapid and sensitive quantification of leading pathogens Cryptosporidium parvum, Giardia gamblia, C ylobacter jejuni, and Escherichia coli from tap water s les with volumes up to 100 mL, and the potential to scale up to larger volumes. qPCR limits of quantification as low as four oocysts for Cryptosporidium, twelve cysts for Giardia, two cells for C. jejuni, and nineteen cells for E. coli per reaction were achieved. A polycarbonate filter-based s ling method coupled with the prepGEM enzyme-based DNA extraction system created a single-step transfer workflow that required as little as 20 min of incubation time and a 100 µL reaction mix. The quantification via qPCR was performed directly on the prepGEM extract, bypassing time-consuming, labour-intensive conventional culture-based methods. The tap water s les were shown to contain insoluble particles that inhibited detection by reducing the quantification efficiency of a representative pathogen (C. jejuni) to 30–60%. This s le inhibition was effectively removed by an on-filter treatment of 20% (v/v) phosphoric acid wash. Overall, the established workflow was able to achieve quantification efficiencies of 92% and higher for all four leading water pathogens, forming the basis of a rapid, portable, and low-cost solution to water monitoring.

Engineering protein nanocages for targeted photodynamic therapy

Publisher: Elsevier BV

Date: 10-2018

DOI: 10.1016/J.NBT.2018.05.171

Development of screening strategies for the identification of paramylon-degrading enzymes

Publisher: Oxford University Press (OUP)

Date: 06-2019

DOI: 10.1007/S10295-019-02157-7

Abstract: Enzymatic degradation of the β-1,3-glucan paramylon could enable the production of bioactive compounds for healthcare and renewable substrates for biofuels. However, few enzymes have been found to degrade paramylon efficiently and their enzymatic mechanisms remain poorly understood. Thus, the aim of this work was to find paramylon-degrading enzymes and ways to facilitate their identification. Towards this end, a Euglena gracilis-derived cDNA expression library was generated and introduced into Escherichia coli. A flow cytometry-based screening assay was developed to identify E. gracilis enzymes that could hydrolyse the fluorogenic substrate fluorescein di-β-d-glucopyranoside in combination with time-saving auto-induction medium. In parallel, four amino acid sequences of potential E. gracilis β-1,3-glucanases were identified from proteomic data. The open reading frame encoding one of these candidate sequences (light_m.20624) was heterologously expressed in E. coli. Finally, a Congo Red dye plate assay was developed for the screening of enzyme preparations potentially able to degrade paramylon. This assay was validated with enzymes assumed to have paramylon-degrading activity and then used to identify four commercial preparations with previously unknown paramylon degradation ability.

Nuclear transformation of the versatile microalga Euglena gracilis

Publisher: Elsevier BV

Date: 2019

DOI: 10.1016/J.ALGAL.2018.11.022

Bioremediation of industrial pollutants by insects expressing a fungal laccase

Publisher: Cold Spring Harbor Laboratory

Date: 28-04-2021

DOI: 10.1101/2021.04.28.441736

Abstract: Inadequate management of household and industrial wastes pose major challenges to human and environmental health. Advances in synthetic biology may help address these challenges by engineering biological systems to perform new functions such as biomanufacturing of high-value compounds from low-value waste streams and bioremediation of industrial pollutants. The current emphasis on microbial systems for biomanufacturing, which often require highly pre-processed inputs and sophisticated infrastructure, is not feasible for many waste streams. Concerns about transgene biocontainment have limited the release of engineered microbes or plants for bioremediation. Engineering animals may provide opportunities for utilizing various waste streams that are not suitable for microbial biomanufacturing while effective transgene biocontainment options should enable in situ bioremediation. Here, we engineer the model insect Drosophila melanogaster to express a functional laccase from the fungus Trametes trogii . Laccase expressing flies reduced concentrations of the endocrine disruptor bisphenol A by more than 50% when present in their growth media. A lyophilized powder made from engineered adult flies retained substantial enzymatic activity, degrading more than 90% of bisphenol A and the textile dye indigo carmine in aqueous solutions. Our results demonstrate that transgenic animals may be used to bioremediate environmental contaminants in vivo and serve as novel production platforms for industrial enzymes. These results support further development of insects, and possibly other animals, as bioproduction platforms and their potential use in bioremediation.

Photoluminescent nanoconjugates for molecular imaging of bladder cancer

Publisher: American Association for Cancer Research (AACR)

Date: 08-2020

DOI: 10.1158/1557-3265.BLADDER19-B27

Abstract: Introduction and Objectives: Despite multiple resections and long-term chemo and immunotherapy, most non-muscle invasive bladder cancer patients suffer from recurrence or progression leading to cystectomy and a less favorable outcome. Possible reasons for that are incomplete resection and reimplantation of cancer cells, which could be prevented by improved resection and adjuvant therapy. Our objective was to develop a targeted drug for detection, fluorescence-guided resection, and deep-penetrating adjuvant photodynamic therapy of urothelial carcinoma (UC). The agent was based on upconversion nanoparticles (UCNP), which can carry a photosensitizer and can transform deep-penetrating near-infrared light into high-energy visible light, demanded for tumor visualization and for production of reactive oxygen species in the photosensitizer. At this stage, we aimed to select an antibody that could be attached to UCNPs to deliver them to UC cells. Methods: We produced silica-coated UCNP of the composition NaYF4:Yb,Er amenable for conjugation with biomolecules. An anti-Glypican-1 (GPC-1) monoclonal antibody MIL-38 (Glytherix Ltd., Sydney, Australia), was chosen for targeted delivery of the nanoparticles as it had previously demonstrated affinity towards bladder cancer. UCNPs were conjugated with MIL-38 by using a fusion protein Linker-Protein G (LPG). Finally, to investigate targeted binding and molecular specificity of these nanoconjugates, we incubated them with GPC-1 positive and GPC-1 negative cells. The role of MIL-38 in targeted delivery of nanoconjugates was also validated by incubation of GPC-1 positive T24 cells with nanoparticles coupled to an isotype control antibody and without an antibody. Results: Targeted upconversion nanoconjugates UCNP-LPG-MIL-38 labeled almost 90% of T24 cells with high expression of GPC-1 and only 23.2% of C3 cells with low expression of this antigen, demonstrating high molecular selectivity and specificity. Incubation of T24 cells with nanoconjugates linked with a control antibody and without antibody resulted in labeling of 19.8% and 26.2%, respectively, demonstrating the role of MIL-38 in targeted delivery of these nanoconjugates. As a result of the labeling, mean photoluminescence of cells in targeted group was from five to eight times stronger than in control groups, allowing for easy identification of positive cells with low background autofluorescence. Conclusions: These results highlight the potential of these nanoconjugates for the diagnosis and therapy of UC, as they can bind to Glypican-1-positive BC cells and cause their bright photoluminescence, which could be used for detection of tumors and activation of photosensitizers. It was also confirmed that monoclonal antibody MIL-38 has high potential to be applied in experimental diagnosis, drug delivery, and targeted therapy of UC, as it mediated targeted binding of upconversion photoluminescent nanoconjugates to Glypican-1-positive UC cells. Citation Format: Liuen Liang, Andrew Care, Anwar Sunna, Douglas C bell, Bradley Walsh, Andrei Zvyagin, David Gillatt, Dmitry Polikarpov. Photoluminescent nanoconjugates for molecular imaging of bladder cancer [abstract]. In: Proceedings of the AACR Special Conference on Bladder Cancer: Transforming the Field 2019 May 18-21 Denver, CO. Philadelphia (PA): AACR Clin Cancer Res 2020 (15_Suppl):Abstract nr B27.

Cell-free enzymatic conversion of spent coffee grounds into the platform chemical lactic acid

Publisher: Frontiers Media SA

Date: 03-12-2019

DOI: 10.3389/FBIOE.2019.00389

Experimental and theoretical tools to elucidate the binding mechanisms of solid-binding peptides

Publisher: Elsevier BV

Date: 09-2019

DOI: 10.1016/J.NBT.2019.04.001

Abstract: The interactions between biomolecules and solid surfaces play an important role in designing new materials and applications which mimic nature. Recently, solid-binding peptides (SBPs) have emerged as potential molecular building blocks in nanobiotechnology. SBPs exhibit high selectivity and binding affinity towards a wide range of inorganic and organic materials. Although these peptides have been widely used in various applications, there is a need to understand the interaction mechanism between the peptide and its material substrate, which is challenging both experimentally and theoretically. This review describes the main characterisation techniques currently available to study SBP-surface interactions and their contribution to gain a better insight for designing new peptides for tailored binding.

Bioremediation of Industrial Pollutants by Insects Expressing a Fungal Laccase

Publisher: American Chemical Society (ACS)

Date: 09-12-2021

DOI: 10.1021/ACSSYNBIO.1C00427

Abstract: Inadequate management of household and industrial wastes poses major challenges to human and environmental health. Advances in synthetic biology may help address these challenges by engineering biological systems to perform new functions such as biomanufacturing of high-value compounds from low-value waste streams and bioremediation of industrial pollutants. The current emphasis on microbial systems for biomanufacturing, which often requires highly preprocessed inputs and sophisticated infrastructure, is not feasible for many waste streams. Furthermore, concerns about transgene biocontainment have limited the release of engineered microbes or plants for bioremediation. Engineering of animals may provide opportunities for utilizing various waste streams that are not suitable for microbial biomanufacturing while effective transgene biocontainment options should enable

Facile Assembly of Functional Upconversion Nanoparticles for Targeted Cancer Imaging and Photodynamic Therapy

Publisher: American Chemical Society (ACS)

Date: 06-05-2016

DOI: 10.1021/ACSAMI.6B00713

Abstract: The treatment depth of existing photodynamic therapy (PDT) is limited because of the absorption of visible excitation light in biological tissue. It can be augmented by means of upconversion nanoparticles (UCNPs) transforming deep-penetrating near-infrared (NIR) light to visible light, exciting PDT drugs. We report here a facile strategy to assemble such PDT nanocomposites functionalized for cancer targeting, based on coating of the UCNPs with a silica layer encapsulating the Rose Bengal photosensitizer and bioconjugation to antibodies through a bifunctional fusion protein consisting of a solid-binding peptide linker genetically fused to Streptococcus Protein G'. The fusion protein (Linker-Protein G) mediates the functionalization of silica-coated UCNPs with cancer cell antibodies, allowing for specific target recognition and delivery. The resulting nanocomposites were shown to target cancer cells specifically, generate intracellular reactive oxygen species under 980 nm excitation, and induce NIR-triggered phototoxicity to suppress cancer cell growth in vitro.

Pseudomonas aeruginosa inhibits the growth of Scedosporium aurantiacum, an opportunistic fungal pathogen isolated from the lungs of cystic fibrosis patients

Publisher: Frontiers Media SA

Date: 24-08-2015

DOI: 10.3389/FMICB.2015.00866

Microwave pretreatment of paramylon enhances the enzymatic production of soluble β-1,3-glucans with immunostimulatory activity

Publisher: Elsevier BV

Date: 09-2018

DOI: 10.1016/J.CARBPOL.2018.05.038

Abstract: A hydrothermal microwave pretreatment was established to facilitate the enzymatic production of soluble bioactive β-1,3-glucans from the recalcitrant substrate paramylon. The efficacy of this pretreatment was monitored with a newly developed direct Congo Red dye-based assay over a range of temperatures. Microwave pretreatment at 170 °C for 2 min resulted in a significantly enhanced enzymatic hydrolysis of paramylon. The action of endo-β-1,3- and exo- β-1,3-glucanases on the microwave-pretreated paramylon produced soluble β-1,3-glucans with degrees of polymerisation (DP) ranging from 2-59 and 2-7, respectively. In comparison, acid-mediated hydrolysis of untreated paramylon resulted in β-1,3-glucans with a DP range of 2-38. The hydrolysates were assayed on their immunostimulatory effect on murine macrophages by measuring the production of the inflammation-linked marker tumour necrosis factor alpha (TNFα) using immunofluorescence. All of the tested hydrolysis products were shown to induce TNFα production, with the most significant immunostimulatory effect observed with the hydrolysate from the exo-β-1,3-glucanase treatment.

Bioproducts From Euglena gracilis

Publisher: Frontiers Media SA

Date: 15-05-2019

DOI: 10.3389/FBIOE.2019.00108

Bioengineering a light-responsive encapsulin nanoreactor: a potential tool for photodynamic therapy

Publisher: Cold Spring Harbor Laboratory

Date: 07-06-2020

DOI: 10.1101/2020.06.06.138305

Abstract: Encapsulins, a prokaryotic class of self-assembling protein nanocompartments, are being re-engineered to serve as ‘nanoreactors’ for the augmentation or creation of key biochemical reactions. However, approaches that allow encapsulin nanoreactors to be functionally activated with spatial and temporal precision is lacking. We report the construction of a light-responsive encapsulin nanoreactor for “on-demand” production of reactive oxygen species (ROS). Herein, encapsulins were loaded with the fluorescent flavoprotein mini-Singlet Oxygen Generator (miniSOG), a biological photosensitizer that is activated by blue-light to generate ROS, primarily singlet oxygen ( 1 O 2 ). We established that the nanocompartments stably encased miniSOG, and in response to blue-light were able to mediate the photoconversion of molecular oxygen into ROS. Using an in vitro model of lung cancer, ROS generated by the nanoreactor was shown to trigger photosensitized oxidation reactions that exerted a toxic effect on tumour cells, suggesting utility in photodynamic therapy. This encapsulin nanoreactor thus represents a platform for the light-controlled initiation and/or modulation of ROS-driven processes in biomedicine and biotechnology.

A platform technology for the bioconjugation of nanoparticles in cancer theranostics

Publisher: Elsevier BV

Date: 10-2018

DOI: 10.1016/J.NBT.2018.05.061

Tools and strategies for constructing cell-free enzyme pathways

Publisher: Elsevier BV

Date: 2019

DOI: 10.1016/J.BIOTECHADV.2018.11.007

Abstract: Single enzyme systems or engineered microbial hosts have been used for decades but the notion of assembling multiple enzymes into cell-free synthetic pathways is a relatively new development. The extensive possibilities that stem from this synthetic concept makes it a fast growing and potentially high impact field for biomanufacturing fine and platform chemicals, pharmaceuticals and biofuels. However, the translation of in idual single enzymatic reactions into cell-free multi-enzyme pathways is not trivial. In reality, the kinetics of an enzyme pathway can be very inadequate and the production of multiple enzymes can impose a great burden on the economics of the process. We examine here strategies for designing synthetic pathways and draw attention to the requirements of substrates, enzymes and cofactor regeneration systems for improving the effectiveness and sustainability of cell-free biocatalysis. In addition, we comment on methods for the immobilisation of members of a multi-enzyme pathway to enhance the viability of the system. Finally, we focus on the recent development of integrative tools such as in silico pathway modelling and high throughput flux analysis with the aim of reinforcing their indispensable role in the future of cell-free biocatalytic pathways for biomanufacturing.

Application of an ELISA-type screen printed electrode-based potentiometric assay to the detection of Cryptosporidium parvum oocysts

Publisher: Elsevier BV

Date: 11-2013

DOI: 10.1016/J.MIMET.2013.08.014

Abstract: We report a novel electrochemical method for the rapid detection of the parasitic protozoan, Cryptosporidium parvum. An antibody-based capture format was transferred onto screen-printed electrodes and the presence of horseradish peroxidase-labelled antibodies binding to the oocysts was potentiometrically detected. This method allowed the detection of 5 × 10(2)Cryptosporidium oocysts per mL in 60 min.

A portable nucleic acid detection system using natural convection combined with a smartphone

Publisher: Elsevier BV

Date: 06-2019

DOI: 10.1016/J.BIOS.2019.03.050

Abstract: The development of portable nucleic acid diagnostic devices has the potential to expand the availability of molecular diagnostics into low-resource settings. One of the promising solutions for rapid and simple DNA lification is the use of Rayleigh-Bernard natural convection which is caused by a buoyancy-driven thermal gradient of liquid when heated from below. This natural convection avoids the use of the complex and sophisticated hardware that is required for precise maintenance of temperature cycles in conventional PCR. We have developed a stand-alone convective PCR (cPCR) device linked to a smartphone for rapid detection of nucleic acids using natural convection heating. The device lifies multiple DNA s les simultaneously using a custom-made heat block controlled by Bluetooth wireless communication. The entire device is highly portable, user-friendly, battery-operated and can provide target DNA lification in less than 30 min. A detection limit of 2.8 × 10

Proteomic response of Euglena gracilis to heavy metal exposure – Identification of key proteins involved in heavy metal tolerance and accumulation

Publisher: Elsevier BV

Date: 2020

DOI: 10.1016/J.ALGAL.2019.101764

Tunable lifetime multiplexing using luminescent nanocrystals

Publisher: Springer Science and Business Media LLC

Date: 15-12-2014

DOI: 10.1038/NPHOTON.2013.322

Solid-Binding Peptides in Biomedicine.

Publisher: Springer International Publishing

Date: 2017

DOI: 10.1007/978-3-319-66095-0_2

Abstract: Some peptides are able to bind to inorganic materials such as silica and gold. Over the past decade, Solid-binding peptides (SBPs) have been used increasingly as molecular building blocks in nanobiotechnology. These peptides show selectivity and bind with high affinity to a erse range of inorganic surfaces e.g. metals, metal oxides, metal compounds, magnetic materials, semiconductors, carbon materials, polymers and minerals. They can be used in applications such as protein purification and synthesis, assembly and the functionalization of nanomaterials. They offer simple and versatile bioconjugation methods that can increase biocompatibility and also direct the immobilization and orientation of nanoscale entities onto solid supports without impeding their functionality. SBPs have been employed in numerous nanobiotechnological applications such as the controlled synthesis of nanomaterials and nanostructures, formation of hybrid biomaterials, immobilization of functional proteins and improved nanomaterial biocompatibility. With advances in nanotechnology, a multitude of novel nanomaterials have been designed and synthesized for diagnostic and therapeutic applications. New approaches have been developed recently to exert a greater control over bioconjugation and eventually, over the optimal and functional display of biomolecules on the surfaces of many types of solid materials. In this chapter we describe SBPs and highlight some selected ex les of their potential applications in biomedicine.

Developing Protein-Based Nanoparticles as Versatile Delivery Systems for Cancer Therapy and Imaging

Publisher: MDPI AG

Date: 16-09-2019

DOI: 10.3390/NANO9091329

Abstract: In recent years, it has become apparent that cancer nanomedicine’s reliance on synthetic nanoparticles as drug delivery systems has resulted in limited clinical outcomes. This is mostly due to a poor understanding of their “bio–nano” interactions. Protein-based nanoparticles (PNPs) are rapidly emerging as versatile vehicles for the delivery of therapeutic and diagnostic agents, offering a potential alternative to synthetic nanoparticles. PNPs are abundant in nature, genetically and chemically modifiable, monodisperse, biocompatible, and biodegradable. To harness their full clinical potential, it is important for PNPs to be accurately designed and engineered. In this review, we outline the recent advancements and applications of PNPs in cancer nanomedicine. We also discuss the future directions for PNP research and what challenges must be overcome to ensure their translation into the clinic.

Multifunctional luminescent nanofibres from Eu3+-doped La2O2SO4 with enhanced oxygen storage capability

Publisher: Elsevier BV

Date: 02-2017

DOI: 10.1016/J.JALLCOM.2016.10.164

Molecular tools and applications of Euglena gracilis: From biorefineries to bioremediation

Publisher: Wiley

Date: 06-08-2020

DOI: 10.1002/BIT.27516

An innovative approach to bioremediation of mercury contaminated soils from industrial mining operations

Publisher: Elsevier BV

Date: 10-2017

DOI: 10.1016/J.CHEMOSPHERE.2017.06.051

Abstract: Soils contaminated with mercury (Hg) have proved expensive and logistically difficult to remediate. Research continues into finding suitable environmentally-friendly and efficient ways of achieving this end. Bioremediation is an option, which employs the strategies microorganisms have evolved to deal with Hg. One microbial strategy involves uptake and intracellular volatilisation of mercuric ions, which passively diffuse from the cell and back into the atmosphere. In this work, Pseudomonas veronii cells grown to stationary phase were immobilised in a xanthan gum-based biopolymer via encapsulation. The P. veronii-biopolymer mix was then coated onto natural zeolite granules. Zeolite immobilised cells remained viable for at least 16 weeks stored under ambient room temperature. Furthermore, the immobilised cells were shown to retain both viability and Hg volatilisation functionality after transportation from Australia to the USA, where they were applied to Hg contaminated soil. Maximum flux rates exceeded 10 μg Hg m

Functionalized Upconversion Nanoparticles for Targeted Labelling of Bladder Cancer Cells

Publisher: MDPI AG

Date: 03-12-2019

DOI: 10.3390/BIOM9120820

Abstract: Bladder cancer is the ninth most common cancer worldwide. Due to a high risk of recurrence and progression of bladder cancer, every patient needs long-term surveillance, which includes regular cystoscopy, sometimes followed by a biopsy of suspicious lesions or resections of recurring tumours. This study addresses the development of novel biohybrid nanocomplexes representing upconversion nanoparticles (UCNP) coupled to antibodies for photoluminescent (PL) detection of bladder cancer cells. Carrying specific antibodies, these nanoconjugates selectively bind to urothelial carcinoma cells and make them visible by emitting visible PL upon excitation with deeply penetrating near-infrared light. UCNP were coated with a silica layer and linked to anti-Glypican-1 antibody MIL38 via silica-specific solid-binding peptide. Conjugates have been shown to specifically attach to urothelial carcinoma cells with high expression of Glypican-1. This result highlights the potential of produced conjugates and conjugation technology for further studies of their application in the tumour detection and fluorescence-guided resection.

Rapid and specific duplex detection of methicillin-resistant Staphylococcus aureus genes by surface-enhanced Raman spectroscopy

Publisher: Royal Society of Chemistry (RSC)

Date: 2020

DOI: 10.1039/C9AN01959F

Abstract: A highly specific method for rapid detection of MRSA genes has been proposed by combining surface-enhanced Raman spectroscopy nanotags and magnetic isolation, which shows great potential for accurate identification of MRSA at an early-diagnosis stage.

Solid-binding peptides for immobilisation of thermostable enzymes to hydrolyse biomass polysaccharides.

Publisher: Springer Science and Business Media LLC

Date: 02-02-2017

DOI: 10.1186/S13068-017-0715-2

Industrial Transformation Training Centres - Grant ID: IC130100009

Start Date: 05-2013

End Date: 09-2017

Amount: $2,100,000.00

Funder: Australian Research Council

Linkage Projects - Grant ID: LP140100462

Start Date: 04-2015

End Date: 12-2020

Amount: $370,000.00

Funder: Australian Research Council

Industrial Transformation Training Centres - Grant ID: IC210100040

Start Date: 02-2022

End Date: 02-2027

Amount: $4,997,903.00

Funder: Australian Research Council