Browsing by Author "Liu, Qing"
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- Advanced DNA Nanopore Technologies
A2 Katsausartikkeli tieteellisessä aikakauslehdessä(2020-09-21) Shen, Boxuan; Piskunen, Petteri; Nummelin, Sami; Liu, Qing; Kostiainen, Mauri A.; Linko, VeikkoDiverse nanopore-based technologies have substantially expanded the toolbox for label-free single-molecule sensing and sequencing applications. Biological protein pores, lithographically fabricated solid-state and graphene nanopores, and hybrid pores are in widespread use and have proven to be feasible devices for detecting amino acids, polynucleotides, and their specific conformations. However, despite the indisputable and remarkable advantages in technological exploration and commercialization of such equipment, the commonly used methods may lack modularity and specificity in characterization of particular phenomena or in development of nanopore-based devices. In this review, we discuss DNA nanopore techniques that harness the extreme addressability, precision, and modularity of DNA nanostructures that can be incorporated as customized gates or plugs into for example lipid membranes, solid-state pores, and nanocapillaries, thus forming advanced hybrid instruments. In addition to these, there exist a number of diverse DNA-assisted nanopore-based detection and analysis methods. Here, we introduce different types of DNA nanostructure-based pore designs and their intriguing properties as well as summarize the extensive collection of current and future technologies and applications that can be realized through combining DNA nanotechnology with common nanopore approaches. - Cationic cellulose nanocrystals for fast, efficient and selective heparin recovery
A1 Alkuperäisartikkeli tieteellisessä aikakauslehdessä(2021-09-15) Liu, Qing; Meng, Zhuojun; Korpi, Antti; Kontturi, Eero; Kostiainen, Mauri A.Heparin is one of the most important anticoagulant agents used in clinical applications. Commercial heparin production includes an isolation from mucosa and an additional enrichment step by cationic resins. However, this process remains time-consuming while heparin is obtained in very low concentrations with the presence of macromolecular impurities, such as proteins. Therefore, an alternative with a fast, efficient and selective heparin-recovery performance is highly desirable. In this work, we utilized a biomass-derived cellulose nanocrystal colloid conjugated with cationic polyelectrolytes for heparin recovery. The high specific surface area and brush-like structure significantly increased the heparin-capture speed and efficiency under physiologically relevant conditions, which were demonstrated by the methylene blue binding assay and quartz crystal microbalance measurement. We also found that a selective heparin capture can be realized via adjusting salt concentration or pH. Finally, we showed that after several recycle rounds, the heparin-recovery ability of the cationic nanocrystals was largely retained and the majority of active heparin dose was recovered, showing a significantly higher heparin-recovery performance than the commercial Amberlite IRA-900 and demonstrating its applicability from an economic perspective. Therefore, the reported cellulose nanocrystal-polymer conjugate represents a promising candidate for a green and efficient heparin recovery. - Engineered protein cages for selective heparin encapsulation
A1 Alkuperäisartikkeli tieteellisessä aikakauslehdessä(2021-02-07) Välimäki, Salla; Liu, Qing; Schoonen, Lise; F. M. Vervoort, Daan; Nonappa, Nonappa; Linko, Veikko; Nolte, Roeland J. M.; C. M. van Hest, Jan; Kostiainen, MauriA heparin-specific binding peptide was conjugated to a cowpea chlorotic mottle virus (CCMV) capsid protein, which was subsequently allowed to encapsulate heparin and form capsid-like protein cages. The encapsulation is specific and the capsid-heparin assemblies display negligible hemolytic activity, indicating proper blood compatibility and promising possibilities for heparin antidote applications. - Engineered Protein Copolymers for Heparin Neutralization and Detection
A1 Alkuperäisartikkeli tieteellisessä aikakauslehdessä(2023-02-13) Liu, Qing; Shaukat, Ahmed; Meng, Zhoujun; Nummelin, Sami; Tammelin, Tekla; Kontturi, Eero; De Vries, Renko; Kostiainen, MauriHeparin is a widely applied anticoagulant agent. However, in clinical practice, it is of vital importance to reverse its anticoagulant effect to restore the blood-clotting cascade and circumvent side effects. Inspired by protein cages that can encapsulate and protect their cargo from surroundings, we utilize three designed protein copolymers to sequester heparin into inert nanoparticles. In our design, a silk-like sequence provides cooperativity between proteins, generating a multivalency effect that enhances the heparin-binding ability. Protein copolymers complex heparin into well-defined nanoparticles with diameters below 200 nm. We also develop a competitive fluorescent switch-on assay for heparin detection, with a detection limit of 0.01 IU mL–1 in plasma that is significantly below the therapeutic range (0.2–8 IU mL–1). Moreover, moderate cytocompatibility is demonstrated by in vitro cell studies. Therefore, such engineered protein copolymers present a promising alternative for neutralizing and sensing heparin, but further optimization is required for in vivo applications. - mRNA detection using constrained DNA hybridization chain reaction on nanoparticles
Kemian tekniikan korkeakoulu | Master's thesis(2020-06-15) Sydänmaanlakka, HenriHybridization chain reaction (HCR) is a powerful tool for the detection of RNA with excellent sensitivity. However, the speed and sensitivity of HCR is usually limited due to its reliance on diffusion kinetics. Constraining the chain reaction in close space allows faster detection speeds and higher sensitivity than non-constrained ones because no diffusion process is needed. The aim of this thesis is to improve the RNA detection speed and sensitivity using HCR. To achieve this, two DNA HCR hairpin monomers were used. After confirming the successful HCR in solution, the HCR monomers were constrained on nanoparticles. The attachment of the DNA hairpins on the surface of the nanoparticles were studied via UV/Vis spectroscopy and verified with transmission electron microscopy. The HCR on top of nanoparticles was studied by measuring time-dependent fluorescence intensity. Successful HCR in solution was verified with polyacrylamide gel electrophoresis and fluorescence measurement and successful attachment of hairpin DNA on top of both gold and silicon nanoparticles was demonstrated. However, the DNA hairpins were partially opened after attachment, which induced the HCR in the absence of the initiator strand. This was verified with polyacrylamide gel electrophoresis. Nevertheless, HCR was proven to be feasible on nanoparticles’ surface and the design needs to be optimized before the method is viable for RNA detection in vivo. - Optically Controlled Construction of Three-Dimensional Protein Arrays
A1 Alkuperäisartikkeli tieteellisessä aikakauslehdessä(2023-07-10) Liu, Qing; Zhou, Yu; Shaukat, Ahmed; Meng, Zhuojun; Kyllönen, Daniella; Seitz, Iris; Langerreiter, Daniel; Kuntze, Kim; Priimagi, Arri; Zheng, Lifei; Kostiainen, Mauri A.Protein crystallization is an important tool for structural biology and nanostructure preparation. Here, we report on kinetic pathway-dependent protein crystals that are controlled by light. Photo-responsive crystallites are obtained by complexing the model proteins with cationic azobenzene dyes. The crystalline state is readily switched to a dispersed phase under ultraviolet light and restored by subsequent visible-light illumination. The switching can be reversibly repeated for multiple cycles without noticeable structure deterioration. Importantly, the photo-treatment not only significantly increases the crystallinity, but creates crystallites at conditions where no ordered lattices are observed upon directly mixing the components. Further control over the azobenzene isomerization kinetics produces protein single crystals of up to ≈50 μm. This approach offers an intriguing method to fabricate metamaterials and study optically controlled crystallization. - Polyelectrolyte Encapsulation and Confinement within Protein Cage-Inspired Nanocompartments
A2 Katsausartikkeli tieteellisessä aikakauslehdessä(2021-10) Liu, Qing; Shaukat, Ahmed; Kyllönen, Daniella; Kostiainen, Mauri A.Protein cages are nanocompartments with a well-defined structure and monodisperse size. They are composed of several individual subunits and can be categorized as viral and non-viral protein cages. Native viral cages often exhibit a cationic interior, which binds the anionic nucleic acid genome through electrostatic interactions leading to efficient encapsulation. Non-viral cages can carry various cargo, ranging from small molecules to inorganic nanoparticles. Both cage types can be functionalized at targeted locations through genetic engineering or chemical modification to entrap materials through interactions that are inaccessible to wild-type cages. Moreover, the limited number of constitutional subunits ease the modification efforts, because a single modification on the subunit can lead to multiple functional sites on the cage surface. Increasing efforts have also been dedicated to the assembly of protein cage-mimicking structures or templated protein coatings. This review focuses on native and modified protein cages that have been used to encapsulate and package polyelectrolyte cargos and on the electrostatic interactions that are the driving force for the assembly of such structures. Selective encapsulation can protect the payload from the surroundings, shield the potential toxicity or even enhance the intended performance of the payload, which is appealing in drug or gene delivery and imaging. - Robotic DNA Nanostructures
A2 Katsausartikkeli tieteellisessä aikakauslehdessä(2020-08-21) Nummelin, Sami; Shen, Boxuan; Piskunen, Petteri; Liu, Qing; Kostiainen, Mauri A.; Linko, VeikkoOver the past decade, DNA nanotechnology has spawned a broad variety of functional nanostructures tailored toward the enabled state at which applications are coming increasingly in view. One of the branches of these applications is in synthetic biology, where the intrinsic programmability of the DNA nanostructures may pave the way for smart task-specific molecular robotics. In brief, the synthesis of the user-defined artificial DNA nano-objects is based on employing DNA molecules with custom lengths and sequences as building materials that predictably assemble together by obeying Watson-Crick base pairing rules. The general workflow of creating DNA nanoshapes is getting more and more straightforward, and some objects can be designed automatically from the top down. The versatile DNA nano-objects can serve as synthetic tools at the interface with biology, for example, in therapeutics and diagnostics as dynamic logic-gated nanopills, light-, pH-, and thermally driven devices. Such diverse apparatuses can also serve as optical polarizers, sensors and capsules, autonomous cargo-sorting robots, rotary machines, precision measurement tools, as well as electric and magnetic-field directed robotic arms. In this review, we summarize the recent progress in robotic DNA nanostructures, mechanics, and their various implementations. - Serum Albumin-Peptide Conjugates for Simultaneous Heparin Binding and Detection
A1 Alkuperäisartikkeli tieteellisessä aikakauslehdessä(2019) Liu, Qing; Välimäki, Salla; Shaukat, Ahmed; Shen, Boxuan; Linko, Veikko; Kostiainen, Mauri A.Heparin is a polysaccharide-based anticoagulant agent, which is widely used in surgery and blood transfusion. However, overdosage of heparin may cause severe side effects such as bleeding and low blood platelet count. Currently, there is only one clinically licensed antidote for heparin: Protamine sulfate, which is known to provoke adverse effects. In this work, we present a stable and biocompatible alternative for protamine sulfate that is based on serum albumin, which is conjugated with a variable number of heparin-binding peptides. The heparin-binding efficiency of the conjugates was evaluated with methylene blue displacement assay, dynamic light scattering, and anti-Xa assay. We found that multivalency of the peptides played a key role in the observed heparin-binding affinity and complex formation. The conjugates had low cytotoxicity and low hemolytic activity, indicating excellent biocompatibility. Furthermore, a sensitive DNA competition assay for heparin detection was developed. The detection limit of heparin was 0.1 IU/mL, which is well below its therapeutic range (0.2-0.4 IU/mL). Such biomolecule-based systems are urgently needed for next-generation biocompatible materials capable of simultaneous heparin binding and sensing. - A Synthetic Protocell-Based Heparin Scavenger
A1 Alkuperäisartikkeli tieteellisessä aikakauslehdessä(2023-03-29) Liu, Qing; Yang, Shuo; Seitz, Iris; Pistikou, Anna Maria Makri; de Greef, Tom F.A.; Kostiainen, Mauri A.Heparin is a commonly applied blood anticoagulant agent in clinical use. After treatment, excess heparin needs to be removed to circumvent side effects and recover the blood-clotting cascade. Most existing heparin antidotes rely on direct heparin binding and complexation, yet selective compartmentalization and sequestration of heparin would be beneficial for safety and efficiency. However, such systems have remained elusive. Herein, a semipermeable protein-based microcompartment (proteinosome) is loaded with a highly positively charged chitosan derivative, which can induce electrostatics-driven internalization of anionic guest molecules inside the compartment. Chitosan-loaded proteinosomes are subsequently employed to capture heparin, and an excellent heparin-scavenging performance is demonstrated under physiologically relevant conditions. Both the highly positive scavenger and the polyelectrolyte complex are confined and shielded by the protein compartment in a time-dependent manner. Moreover, selective heparin-scavenging behavior over serum albumin is realized through adjusting the localized scavenger or surrounding salt concentrations at application-relevant circumstances. In vitro studies reveal that the cytotoxicity of the cationic scavenger and the produced polyelectrolyte complex is reduced by protocell shielding. Therefore, the proteinosome-based systems may present a novel polyelectrolyte-scavenging method for biomedical applications.