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Sustainable Hydrometallurgy of Energy-Relevant Critical Minerals from Unconventional Sources: Ex-situ Carbon Mineralization and Thermal Swing Salting-Out Process
No full textThe clean energy transition toward carbon neutrality requires shifting from a ‘fossil fuel–based’ to a ‘metal–based’ energy system. However, limited metal reserves demand the extraction of critical minerals such as lithium (Li), nickel (Ni), cobalt (Co), copper (Cu), and rare earth elements (REEs) from unconventional sources like industrial wastes, tailings, low-grade ores, and brines. These sources pose challenges due to low grades and complex chemistries, leading to higher energy use and CO2 emissions per ton of metal produced, creating a fundamental “metal–energy–CO2” dilemma. This study proposes a new framework that integrates carbon mineralization and resource recovery, reconceptualization of mineral resources by valorizing the waste and gangue materials of unconventional sources as CO2 sequestration media. In this framework, the economic viability of lower-grade deposits and wastes may be enhanced by the added decarbonization value, positioning them as potential future reserves. To address the simultaneous handling of abundant alkaline earth metals and trace critical elements, integrated processes were developed: (i) selective REE recovery during indirect carbon mineralization, (ii) chelation- assisted REE extraction during direct mineralization, and (iii) a thermal swing salting-out (TSSO) process for Li extraction from unconventional brines. The REEs recovery research began with collecting 22 types of industrial wastes and investigation physicochemical properties of slags. Compositional analysis revealed that blast furnace slag (BFS) and electric arc furnace slag (EAFS) contained over 300 ppm REEs and 40 wt% of Ca and Mg contents, and are selected as proper candidates. In sequential extraction and n(H+) regulation experiments, REEs were found encapsulated within host matrices, implying the presence of ‘mineralogical barrier’. However, the modified direct and indirect carbon mineralization effectively liberated REEs without impeding CO2 sequestration. A modified pH- swing using oxalic acid significantly enhanced REE recovery, producing REE-rich solids exceeding 1.9 wt% while sequestering 117 kg CO2 per ton of slag. Chelation-assisted direct carbonation further enabled selective extraction of Mn and heavy REEs over Ca, achieving over 85% extraction efficiency. Surprisingly, REE leachability increased with atomic number, attributed to the slightly higher Kf values of heavy REEs (HREEs) compared to light REEs (LREEs). This study highlights that carbon mineralization can act as ‘REEs liberation methodology’ which can make synergies with REEs recovery process. Next, the Li recovery research was focused on developing novel TSSO process. The objective of TSSO process was extracting Li from unconventional brines by utilizing natural thermal energy sources including diurnal temperature variations and geothermal heat. By using thermoresponsive amine-based solvents (i.e., diisopropylamine (DIPA)) and differences of salt solubility, the TSSO process selectively concentrated Li while rejecting impurities. Application to simulated geothermal brine (consist of Li, Na, K) demonstrated a 2.5-fold increase in Li concentration, elevating it to commercially relevant levels (~500 ppm). In an optimum point, 0% of Ca, 1.1% of Na, 1.1% of K, and 52% of Li were recovered in regenerated aqueous solution. These results highlight TSSO process could be used as thermally driven Li extraction method suitable for resource-constrained environments such as alpine and desert regions. In conclusion, this study highlights that a holistic and integrated approach, rather than sector-specific methods, provides an effective strategy for overcoming the interconnected “Metal–Energy–CO2” dilemma. Future advancements toward carbon neutrality and the energy transition will require the development of integrated process frameworks that bridge mining, metallurgy, waste management, carbon sequestration, energy systems, construction, and steel industries.DoctorAbstract i
Contents iii
List of Figures v
List of Tables ix
Chapter 1. Introduction 1
1.1. The Great Acceleration and Linear Economy 1
1.2. A Paradox of Energy Transition: Do We Have Enough Metals 4
1.3. Reconceptualizing Mineral Resources: Unlocking New Values of Unconventional Sources for Critical
Mineral Recovery and CO2 Sequestration 7
1.4. Research Gap: Addressing Dual Objectives and Heterogeneous Metallic Properties from Single
Feedstock 9
1.5. Process System Designation and Tailoring Reaction Pathways of Carbon Mineralization 10
1.6. Research Goal and Outline of the Dissertation 11
Chapter 2. Expanded Concept of Mineral Resource: Systematic Review 14
2.1. Can CO2 Sequestration Capacity Be a Resource 14
2.2. Expanded Concept of Mineral Resource 16
2.3. CO2 Sequestration Potential of Alkaline Industrial Wastes and Natural Minerals 18
2.4. Critical Mineral Concentration in Unconventional Sources 23
Chapter 3. Theories of Carbon Mineralization and Thermal Swing Salting Out Process 35
3.1. Theory of ex-situ Carbon Mineralization and State of Art 35
3.2. Theory of Salting Out and Temperature Swing Solvent Extraction Process 39
Chapter 4. Exploring the Potential of Iron and Steelmaking Slags as Alternative Sources of Rare Earth
Elements and Carbon Mineralization. 42
4.1. Background 42
4.2. Experimental 44
4.3. Results and Discussions 47
4.4. Conclusions 50
Chapter 5. Indirect and Direct Carbon Mineralization with Simultaneous Recovery of Rare Earth
Elements from Industrial Wastes 65
5.1. Background 65
5.2. Thermodynamic Simulations and Hypotheses 66
5.3. Experimental 70
5.3. Results and Discussions 74
5.3.1. Indirect Carbon Mineralization and REEs Recovery 74
5.3.1. Chelation-assisted Direct Carbon Mineralization and REEs Recovery 90
5.4. Conclusions 92
Chapter 6. Alkali Fusion Pretreatment of Blast Furnace Slag for Enhancing Leaching Efficiency of
Metals 98
6.1. Background 98
6.2. Experimental 99
6.3. Results and Discussions. 102
6.3.1. Physicochemical properties of BFS and formation of Si-passivation layer 102
6.3.2. Structural Modifications by Alkali Fusion and Water Leaching Pretreatment 105
6.3.3. Enhanced leaching by alkali fusion-water leaching pretreatment 107
6.4. Conclusions 110
Chapter 7. Novel Application of Thermal Swing Salting-Out Process for Lithium Extraction from
Geothermal Brine 122
7.1. Background 122
7.2. Experimental 124
7.3. Results and Discussions. 128
7.3.1. Preliminary Test and Metal Behaviors of Three-Phase Separation of TSSO 128
7.3.2. Evaluation of process parameters for two-phase separation of TSSO 129
7.3.3. Statistical analysis and simulation of TSSO process 131
7.3.4. Application of TSSO Process for Geothermal Brine 134
7.4. Conclusions 135
Chapter 8. Conclusions and Recommendations for Future Works 153
8.1. Summary 153
8.2. Conclusions 156
8.3. Recommendations for Future Works 157
References 16
Interface engineering for highly efficient and stable perovskite solar cells and their applications
No full textInterface engineering is crucial for achieving highly efficient and stable perovskite solar cells (PSCs). Here, we present a unified strategy that employs organic passivation layers at interfaces in both n–i–p and p–i–n device configurations.
In the n–i–p configurations, a newly synthesized bathocuproine-based polyelectrolyte (poly-BCP) was introduced between the SnO₂ electron transport layer (ETL) and the perovskite absorber. This interlayer effectively passivated oxygen-vacancy defects at the SnO₂ side while simultaneously scavenging ionic defects at the perovskite side, thereby suppressing both bulk and interfacial nonradiative recombination. As a result, the modified PSCs achieved a power conversion efficiency (PCE) of 24.4% with an open-circuit voltage of 1.21 V, and retained 93% of their initial efficiency after 700 hours of continuous one-sun irradiation under inert conditions without encapsulation.
For the p–i–n configuration, amine-functionalized organic small molecules (e.g., PBN) were employed as passivation layers at the interface between the PCBM electron transport layer and the copper electrode. Strong Cu–N coordination facilitated the dense growth of Cu electrodes with strong adhesion, effectively blocking moisture ingress. Consequently, non-encapsulated devices retained 90% of their initial PCE after 200 days under ambient air (25 °C, 20–40% relative humidity), and showed negligible degradation even under harsher conditions such as high humidity or direct water immersion.
Furthermore, we extended this approach to quasi-two-dimensional Ruddlesden–Popper perovskites (RPPs), which are inherently more stable but typically less efficient than their 3D counterparts. By applying a multifunctional organic molecule as a surface passivation treatment on RPP films (PEA₂MA₄Pb₅I₁₆, ⟨n⟩ = 5) in p–i–n devices, both surface and deep-level traps were effectively suppressed through electrostatic interactions, while preferred out-of-plane crystal orientation was induced. These effects facilitated efficient carrier transport, leading to a PCE of 20.05% with negligible hysteresis. The optimized devices maintained 88% of their initial efficiency after 1000 hours of continuous one-sun operation under maximum power point tracking, without encapsulation or UV filtering
What Links Sleep and Neuropathic Pain?: A Literature Review on the Neural Circuits for Sleep and Pain Control
Full text linkNeuropathic pain (NP), arising from lesions or diseases of the sensory nervous system, significantly disrupts sleep, creating a feedback loop where pain and sleep disturbances exacerbate each other. Research suggests that sleep disruption may contribute to progressing from acute to chronic NP. The neural circuits involved in sleep-wake regulation and pain processing are intricately interconnected, making it difficult to pinpoint the specific regions responsible for each function. This review seeks to disentangle these complex relationships by providing a detailed overview of the anatomical pathways involved in NP, extending from the peripheral to the central nervous system. Additionally, it examines the neurocircuits that govern sleep-wake cycles and their interaction with pain pathways. By illuminating these connections, this review aims to deepen our understanding of how sleep influences NP, ultimately guiding the development of more effective strategies for managing NP and its associated sleep disturbances to improve the quality of life for those affected.TRUEsciescopu
Engineering Magnetic Nanobiocatalysts via Mussel Foot Protein-Mediated Laccase Immobilization for Enhanced Performance and Reusability
No full textRobust, reusable, and environmentally benign enzyme-based catalysts are promising for advancing more sustainable chemical processes. Laccase, a multicopper oxidoreductase, can transform a diverse range of phenolic and aromatic pollutants in water; however, limitations in operational stability and reusability hinder application scalability. Here, we report a bioinspired immobilization strategy using microbially derived mussel foot protein-5 (MFP) to anchor laccase onto superparamagnetic iron oxide nanoparticles (IONPs, similar to 8 nm). MFP's high DOPA content, strong metal oxide affinity, and cationic nature enable multipoint electrostatic binding to laccase (pI approximate to 3.5), achieving up to 98% immobilization yield with 80-110% activity recovery, surpassing cationic surfactant ligands and far exceeding anionic or silica coatings (50% lower thermal deactivation at 45 degrees C, and a 20 kJ mol(-1) higher activation energy, reflecting enhanced stability. The system retained 71.9 +/- 8.7% activity after eight reuse cycles and achieved similar to 20% greater methyl orange removal than free laccase. Taken together, this bioinspired, magnetically recoverable platform provides a scalable route to high-efficiency, reusable, and thermodynamically stable oxidoreductase systems for a wide range of catalytic applications.FALSEsciescopu
Piezo-resistive Tactile Sensing System with Artificial Intelligence for User Movement and Intention Recognition
No full textHuman activity recognition plays an important role in the daily life of people because of its ability to learn extensive high-level information about human activity. With the advancement of deep learning, research in this area has been rapidly expanding. However, although understanding the forces and dynamics of movement is crucial for understanding human motion, most studies focus on kinematics rather than kinetics. Tactile sensors can be a solution to these problems. A tactile sensor can measure applied pressure and stress, similar to human skin. Therefore, it enables force-based monitoring of physiological signals and daily physical interactions to human recognition. Among the various types of tactile sensors, the piezo-resistive film is known as a more accessi- ble tactile sensor than others since it is cost-effective and can be manufactured in various shapes. However, recent research on piezo-resistive film-based tactile sensor has primarily aimed at object classification and enhancing robotic grasping capabilities. Studies focused on acquiring and utilizing human movement and intention-related information remain in the early stages. Therefore, in this thesis, we conducted research on Piezo-resistive Tactile Sensing System with Artificial Intelligence for User Movement and Intention Recognition. This doctoral dissertation aims to demonstrate the following key contributions. First, it shows that motion recognition using tactile sensors provides richer information about movement and enables a deeper understanding of human motion compared to conventional pose-based methods. Second, it demonstrates that continuous tracking of human motion using tactile sensors combined with artificial intelligence has the potential to interpret user intent and apply it to human-computer interaction (HCI) and human-robot interaction (HRI). To achieve this, the study presents a system utilizing a high-resolution piezo-resistive film, which allows users to compare their own movements with those of experts during exercise feedback, enabling them to adjust and refine their performance. Furthermore, the research proposes and validates a method for recognizing user motion and intent using both mat-type and custom high-resolution tactile sensors in combination with artificial intelligence. These capabilities are then applied to HCI and HRI scenarios. Through this process, the dissertation demonstrates that high-resolution tactile sensors based on piezo-resistive films are effective in understanding human movement. Moreover, by integrating these sensors with artificial intelligence, the system can recognize and interpret more complex human motions and intentions, extending its applicability across various interactive domains.|사람의 활동 인식(Human Activity Recognition, HAR)은 인간 활동에 대한 고차원 정보를 학습할 수 있다는 점에서 일상생활에서 중요한 역할을 한다. 딥러닝의 발전과 함께 이 분야의 연구는 빠르게 확장되고 있다. 그러나 인간의 움직임을 이해하기 위해서는 힘과 동역학에 대한 이해가 필수적임에도 불구하고, 대부분의 기존 연구는 운동학(kinematics)에 집중하고 있으며 운동역학(kinetics)은 상대적으 로 소홀히 다루어지고 있다. 이러한 한계를 극복할 수 있는 방법 중 하나로 촉각 센서(tactile sensor) 가 제안될 수 있다. 촉각 센서는 인간의 피부처럼 압력과 응력을 측정할 수 있어, 생리 신호나 일상적인 물리적 상호작용을 힘 기반으로 모니터링하고 이를 통해 사람을 인식할 수 있게 해준다. 다양한 유형의 촉각 센서 중에서도, 압저항 필름(piezo-resistive film)은 저비용이며 다양한 형태로 제작이가능하다는점에서접근성이높은센서로알려져있다. 그러나최근의압저항필름기반촉각센서 연구는 대부분 물체 분류나 로봇의 파지(grasping) 성능 향상에 집중되어 왔다. 인간의 움직임 및 의도와 관련된 정보를 수집하고 이를 활용하는 연구는 아직 초기 단계에 머물러 있다. 따라서 본 학위논문에서는 사용자 움직임 및 의도 인식을 위한 인공지능 기반 압저항 촉각 센서 시스템에 대해 연구를 수행하였다. 이 박사학위 논문은 다음과 같은 주요 기여를 목표로 한다. 첫째, 촉각 센서를 활용한 모션 인식이 기존의 자세 기반 방법보다 더 풍부한 움직임 정보를 제공하며 인간 움직임에 대한 더 깊은 이해를 가능 하게 함을 보인다. 둘째, 촉각 센서와 인공지능을 결합하여 인간의 움직임을 연속적으로 추적함으로써 사용자의도를해석하고,이를인간-컴퓨터상호작용(HCI)및인간-로봇상호작용(HRI)에적용할수있는 가능성을 입증한다. 이를 위해 본 연구는 고해상도 압저항 필름을 활용한 시스템을 제안하고, 사용자가 운동 피드백 과정에서 자신의 움직임을 전문가의 움직임과 비교하여 스스로의 운동을 수정해 나갈 수 있는 기능을 구현하고 이를 입증하였다. 또한, 매트형 및 커스터마이징된 부착형 고해상도 촉각 센서를 인공지능과 결합하여 사용자 모션 및 의도를 인식하는 방법을 제안하고, 이를 HCI 및 HRI 시나리오에 적용하여 성능을 검증하였다. 이 과정을 통해 본 논문은 압저항 필름 기반의 고해상도 촉각 센서가 인간 움직임 이해에 효과적임을 보여주며, 나아가 이를 인공지능과 결합함으로써 보다 복잡한 인간의 모션과 의도까지 인식하고 활용할 수 있는 시스템 구현 가능성을 제시한다. 이로써 다양한 상호작용 환경에서의 적용 가능성을 확장한다.DoctorAcknowledgments i
Abstract ii
Contents v
List of Tables ix
List of Figures x
Chapter 1. Introduction 1
Chapter 2. Research 1. Real-time Multi-modal Comprehensive Body-
weight Exercise Feedback System with a Personal Virtual
Expert Guiding 5
2.1 Introduction 5
2.2 Related works 7
2.2.1 Pose feedback techniques 7
2.2.2 Pressure feedback techniques 8
2.3 Exercise feedback methods 8
2.4 System Features 10
2.5 Exercise sensing setup 10
2.6 Multimodal exercise feedback technique 11
2.7 Experimental Setup 16
2.7.1 Research questions 16
v
2.7.2 Experimental conditions 16
2.7.3 Participants 17
2.7.4 Procedure 17
2.7.5 Measurements 18
2.8 Result 21
2.9 Discussion 24
2.10 Limitations and Future Work 28
2.11 Conclusion 28
Chapter 3. Research 2. Seamless-walk: Natural and Comfortable
Virtual Reality Locomotion Method with a High-resolution
Tactile Sensor 30
3.1 Introduction 30
3.2 Related work 32
3.2.1 VR locomotion 32
3.2.2 Foot-based locomotion 33
3.3 Method 36
3.3.1 High resolution tactile sensor 36
3.3.2 Speed and angle prediction 37
3.3.3 Algorithm validity 40
3.4 Experiment 42
3.4.1 Sub experiment1: comparing with hand-based equip-
ment 42
3.4.2 Sub experiment2: comparing with foot-based equip-
ment 46
– vi –
3.4.3 Procedure 48
3.5 Result and discussion 49
3.5.1 Sub experiment1: comparing with hand-based equip-
ment 49
3.5.2 Result of sub experiment2: comparing with foot-based
equipment 54
3.6 Summary 62
Chapter 4. Research 3. Adaptive Walker: User Intention and Ter-
rain Aware Intelligent Walker with High-resolution Tac-
tile and IMU Sensor 63
4.1 INTRODUCTION 63
4.2 Walker System Overview 66
4.2.1 Tactile sensor 67
4.2.2 IMU sensor 68
4.2.3 Computing system 68
4.2.4 Actuation system 69
4.3 User Intention and terrain recognition 70
4.3.1 Intent recognition 71
4.3.2 Terrain recognition 71
4.4 Adaptive walker system integration 73
4.5 System evaluation 74
4.6 Limitations and future works 76
4.7 Conclusion 77
– vii –
Chapter 5. Discussion 79
Chapter 6. Limitations 82
Chapter 7. Future works 83
Chapter 8. Conclusion 84
References 85
Curriculum Vitae 100
– viii
Structural and electrical transitions in VO2/c-Al2O3 films investigated using simultaneous X-ray reciprocal space mapping and electrical resistance measurements
No full textWe carried out simultaneous in-situ three-dimensional X-ray reciprocal space mapping and electrical resistance measurements of VO2 films grown on c-cut sapphire to investigate the relationship between the structural phase transition (SPT) and the metal-insulator transition (MIT). The decoupling of the MIT and SPT behaviors was more pronounced in the thinnest film (37 nm), with a difference in transition temperatures (triangle Tc) of approximately 8.3 degrees C. Despite a decrease of over 50% in the M1 fraction, the electrical resistance remained in the insulating phase, indicating a delay in the MIT. This behavior is attributed to the disconnected island morphology, which restricts the formation of continuous conduction pathways. As the film thickness increased, triangle Tcgradually decreased, and eventually both thermal-hysteresis characteristics became similar at a thickness of 360 nm. Furthermore, pronounced six-fold diffuse X-ray scattering was observed around the VO2 M1 (020) Bragg peak, revealing the presence of structural defects and small crystalline domains. This hexagonal pattern originates from three in-plane variants that are rotated by 120 degrees about the film-normal b-axis. Each variant is paired with a 21 screw-axis twin domain, corresponding to a 180 degrees rotation about the same axis. As a result of this combined rotational and twinning symmetry, the diffuse scattering appears along the a* and c* directions of the M1 phase. The correlation length (xi x) of the monoclinic order steadily decreased even in the pre-transition region, exhibiting a trend similar to that of the electrical resistance. When xi x reached approximately 6 nm, the electrical resistance began to decrease rapidly. This correlated behavior suggests a possible interplay between structural disorder, such as defect generation between coherent domains and limited grain connectivity, and charge transport in VO2 films.FALSEsciescopu
Structural and functional study of transporters: ABCB6 and GLUT7 by using cryo-EM
No full textOver the past decade, remarkable advancements in cryo-electron microscopy (cryo-EM) have
revolutionized the field of structural biology. Among various innovations, my focus has been on the structural analysis of membrane proteins. Due to their intrinsic characteristics, such as requiring diverse membrane reconstitution methods during the purification process, membrane proteins have posed significant challenges in structure determination using X-ray crystallography, which relies on obtaining high-quality crystals for diffraction pattern analysis. However, with the development of cryo-EM, a new avenue has emerged for determining membrane protein structures. Leveraging these advancements, I conducted structural studies on membrane proteins, particularly transporters. In the first part of this study, I targeted ATP Binding Cassette (ABC) transporters, which hydrolyze ATP and exhibit a broad substrate spectrum. Specifically, I determined the structure of ABCB6, the sixth protein in the B family of ABC transporters. Despite ABCB6 being relatively well-studied, I elucidated the previously unresolved transition state, the outward-facing open conformation, at a resolution of 3.8 Å. Additionally, I revealed that lipids play a structural role in stabilizing the protein. In the second part, I determined the structure of glucose transporter 7 (GLUT7), a protein known to transport glucose, one of the most fundamental energy sources in biological systems. Given its small molecular weight of 55 kDa, structure determination using cryo-EM was considered challenging. However, by employing nanodiscs and implementing a reference-based filtering process during data processing to eliminate bad particles, I achieved a final resolution of 3.3 Å. This study provides insights into the structural analysis of membrane proteins and the characterization of protein transition states using cryo-EM. The newly determined structures serve as foundational data for future research in this field.Doctorby employing nanodiscs and implementing a reference-based filtering process during data processing to
eliminate bad particles, I achieved a final resoluti n of 3.3 Å.
This study provides insights into the structural analysis of membrane proteins and the characterization
of protein transition states using cryo-EM. The newly determined structures serve as foundational data for
future research in this field.
CONTENTS
PART I
ABSTRACTS i
CONTENTS iii
LIST OF TABLES AND FIGURES iv
I. INTRODUCTION 1
II. MATERIALS AND METHODS
2.1. Cloning, expression and purification of ABCB6 and its mutants 3
2.2. Cloning, expression and purification of hApoA-I ((Δ1−43) 4
2.3. Nanodisc reconstitution 4
2.4. ATP hydrolysis assay 5
2.5. Proteoliposome reconstitution 5
2.6. Proteoliposome transport activity assay for CPIII 6
2.7. CryoEM grid preparation and data collection 6
2.8 CryoEM data processing 6
2.9 Model building and refinement 7
III. RESULTS
3.1. VO4 and AlF4 can inhibit the catalytic cycle, capturing the post-ATP hydrolysis state 8
3.2. An application to prefer the outward-facing conformation in hABCB6core 8
3.3. The impact of the W546A mutation on the ATPase and transport functions of hABCB6core 9
3.4. The W546A mutant favors the outward-facing state in detergent environment but not in nanodisc
reconstitution · · · · · · · · · · · · · ·· · · · · · · · · · · · · · · · · ·· · · · · · · · ·· · · · · · · · · · · · · · · · · ·· · · · · · · 10
3.5. Overall structure of outward-facing conformation 12
3.6. The outward-facing conformation has low substrate affinity, facilitating its release 12
IV. DISCUSSION 14
LIST OF TABLES AND FIGURES
Figure 1. Inhibition of the ATPase activity of hABC6core by ATP analogs 16
Figure 2. Purification profile of hABCB6coreand proteoliposome 18
Figure 3. Representative cryoEM data processing results 20
Figure 4. Structural difference depending on conformation 22
Figure 5. Well-conserved the central interactions through the substrate pathway ·· 24
Figure 6. ATPase activity of ABCB6core and W546A proteins 26
Figure 7. Purification profile of W546A mutant· 28
Figure 8. Comparison of the hABCB6core - W546A mutant in detergent and Nanodisc 30
Figure 9. Electron density of lipids interacting with TMDs hydrophobic region 32
Figure 10. The post-occluded conformation of the hABCB6core-W546A mutant is
stabilized by a phospholipid embedded within the hydrophobic groove of the
TMD surface 34
Figure 11. The structural overview of the hABCB6core-W546A mutant in an outward-
facing conformation· 36
Figure 12. During the CPIII transport cycle, assumed conformational change on the
substrate binding site with all structure data of ABCB6 38
Figure 13. During the Hemin:GSH transport cycle, assumed conformational change on
the substrate binding site with all structure data of ABCB6· 40
Figure 14. Cryo-EM data processing pipeline employed for detergent-purified
hABCB6core in the presence of CPIII and Mg2+/ADP·VO4 42
Figure 15. Cryo-EM data processing pipeline employed for Mg2+/ADP·VO4-bound,
outward-facing hABCB6core-W546A in detergent micelles 44
Figure 16. Cryo-EM maps of outward-facing hABCB6core-W546A in complex with
Mg2+
/ADP·VO4 46
Figure 17. Cryo-EM data quality analysis of outward-facing hABC 6core-W546A in
complex with Mg2+/ADP·VO4· 48
Figure 18. Cryo-EM data processing pipeline employed for Mg2+/ADP·VO4-bound, post-
occluded hABCB6core-W546A in nanodiscs 50
Figure 19. Cryo-EM maps of the post-occluded hABCB6core-W546A in complex with
Mg2+
/ADP·VO4 52
Figure 20. Cryo-EM data quality analysis of the post-occluded hABCB6core-W546A in
complex with Mg2+
/ADP·VO4 54
Figure 21. Schematic diagrams of conformational equilibria between the outward-facing
and post-occluded configurations of hABCB6core and the W546A mutant
56
Figure 22. Role of conserved aromatic residues in stabilizing the occluded conformation
of the ABC transporter family 58
Table 1. Cryo-EM data collection, refinement and validation statistics 60
PART II
ABSTRACTS i
CONTENTS iii
LIST OF TABLES AND FIGURES iv
I. INTRODUCTION 62
II. MATERIALS AND METHODS
2.1. Cloning, expression and purification 63
2.2. Nanodisc reconstitution 64
2.3. Grid preparation and data acquisition 64
2.4. Electron microscopy data processing 64
2.5. Model building and refinement 65
III. RESULTS AND DICUSSION
3.1. Structure detemination·· ·· · ·· · · ·· · ·· · ·· · ·· · ·· · · ·· · ·· · · ·· · ·· · · ·· · ·· · ·· · ·· · ·· · ·· · · ·· 66
3.2. Overall fold· · · · · · · · · · · · · · · · · · · · · · · · · · · ·· · · · · · · · · ·· · · · ·· · · · · · · · · · ·· · · · · · · · · ·· · · · · 67
3.3. Substrate-binding cavity· 67
3.4. Alternating access cycle· · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · · ·· · · · · 68
LIST OF TABLES AND FIGURES
Figure 1. Cryo-EM image and processing flowchart· 70
Figure 2. The quality of Cryo-EM processing results 72
Figure 3. Topology of human GLUT7 74
Figure 4. Outward-open structure of human GLUT7 76
Figure 5. Substrate-binding cavity 78
Figure 6. Possible mechanism of the conformational transition 80
Table 1. Cryo-EM data collection, refinement and validation statistics 82
ABSTRACT IN KOREAN 85
ACKNOWLEDGEMENT 86
REFERENCES 8
Therapeutic Evaluation of Subcutaneously Delivered, Alginate-Encapsulated, Genetically Engineered Mesenchymal Stem Cells for Fabry disease
No full textFabry disease is an X-linked lysosomal storage disorder caused by α-galactosidase A (α-Gal A) deficiency, leading to globotriaosylceramide accumulation. Current enzyme replacement therapy (ERT) requires biweekly intravenous infusions, presenting significant limitations in patient compliance and quality of life. In this study, we developed a long-term delivery system using genetically engineered mesenchymal stem cells (eMSCs) encapsulated in alginate microgels. Human α-galactosidase A-expressing eMSCs maintained high viability and stable enzyme secretion after encapsulation, and subcutaneous injection of these αGal-eMSC-Alg constructs demonstrated structural stability for up to 12 weeks in nude mice. Therapeutic efficacy in GLA knockout Fabry mice showed age-dependent outcomes, with the aged group exhibiting better trends of eMSC survival and reduced kidney Gb3 accumulation compared to the young group. This αGal-eMSC-alginate system offers single-injection, long-term therapeutic delivery advantages over conventional enzyme replacement therapy, representing a promising platform for lysosomal storage disorders with potential for optimization through further investigation of enhanced immune evasion strategies.MasterChapter 1. Introduction 1
1.1. Fabry Disease and Current Therapeutic Limitations 1
1.2. MSCs as a Platform for Cell-Based Gene Therapy 1
1.3. Alginate Hydrogel Encapsulation for Enhanced Cell Delivery 2
1.4. Experimental overview 3
Chapter 2. Materials and Methods 5
2.1. Materials 5
2.2. Vector Construction and Preparation 5
2.3. in vitro studies 6
2.3.1. Cell culture 6
2.3.2. Plasmid Transfection 7
2.3.3. Lentiviral Transduction 7
2.3.4. Flow Cytometry Analysis 8
2.3.5. Absolute Quantitative PCR 8
2.3.6. α-Galactosidase A Activity Assay 9
2.3.7. Cell Encapsulation in Alginate Microgels 9
2.3.8. Microgel Size Distribution Analysis 10
2.3.9. Live/Dead Viability Assay 10
2.4. in vivo studies 10
2.4.1. Animal Care and Experimentation 10
2.4.2. DiD Labeling and In Vivo Cell Tracking 11
2.4.3. In Vivo Therapeutic Efficacy Study in Fabry Disease Mice 11
2.4.4. Immunofluorescence Staining of Kidney and Gel Tissues 12
2.5. Statistical Analysis 13
Chapter 3. Results and Discussion 14
3.1. Construction of bicistronic Lentiviral Vectors for constant over-secretion of α-Gal A and
GFP expression. 14
3.2. Transient Expression and Secretion of α-Gal A in HEK293T Cells. 14
3.3. Optimization of Lentiviral Transduction Conditions for UC-MSC Engineering. 16
3.4. Enrichment and Validation of Stably Transduced αGal-GFP-eMSCs via Antibiotic
Selection. 18
3.5. Quantification of Lentiviral Vector Copy Number in Genetically Engineered MSCs. 21
3.6. Validation of Cryopreservation Stability and Expression in αGal-eMSCs. 22
3.7. Characterization of αGal-eMSC-Alg: Function and Structural Integrity. 23
3.8. Cell Viability Assessment of αGal-eMSC-Alg at Different Encapsulation Densities. 25
3.9. Long-Term in vivo Retention and Viability of αGal-eMSC-Alg in Nude Mice. 27
3.10. Therapeutic Efficacy Evaluation of αGal-eMSC-Alg in Fabry Disease Mouse Models. 29
3.10.1. Sustained Therapeutic Response in Aged Fabry Disease Mice: Evidence for Long-Term Enzyme Delivery. 31
3.10.2. Transient Response and High Variability in Young Fabry Disease Mice. 34
Chapter 4. Conclusion 38
References 4
PPh2-ImPy Ligands with an Unsymmetric N-Aryl Ring for Efficient Acrylate Synthesis Reaction from Ethylene and CO2
No full textBuilding on the previously reported PPh₂-ImPy ligand system which showed an excellent reactivity in the acrylate synthesis reaction, a new ligand design strategy was applied to further improve reaction efficiency. While maintaining the structural stability of the original ligand system, two substituents with different steric bulkiness were introduced into the N-aryl ring. The resulting unsymmetric steric environment around the Ni center has the possibility that could induce certain catalytic steps to proceed on more or less hindered side depending on their steric demands. This strategy is likely to facilitate ligand dissociation step on the more hindered side, while promoting oxidative coupling with CO2 step on the less hindered side, thereby enabling an efficient progression of the catalytic cycle. As a result, this may lead to enhanced catalytic performance in the acrylate synthesis reaction. Of the four different ligands with an unsymmetric N-aryl ring, the PPh₂–ImPy ligand (TON 146) with i-propyl and n-propyl substituents at the ortho positions of the N-aryl ring showed the highest reactivity in the acrylate synthesis reaction. Also, it exhibited more improved reactivity than the previously reported PPh₂–ImPy ligand (TON 122) with symmetric N-aryl ring. The result showed the possibility that the unsymmetric steric environment tuned by the difference in the steric bulkiness and positional arrangement of each substituent on N-aryl ring could lead to the improvement of the catalytic efficiency in the acrylate synthesis reaction.MasterIntroduction 1
1.1. Carbon Dioxide (CO₂) as a C1 Building Block 1
1.2. Synthesis of Acrylate from Ethylene and CO₂ 2
1.3. Development of Ligand Systems for Acrylate Synthesis 4
1.4. Imidazo[1,5-a]pyridin-3-ylidene (ImPy) Ligand 5
1.5. Application of ImPy to Acrylate Synthesis 7
1.6. Pyridine-Chelated ImPy Ni(II) Complexes 7
1.7. Structural Comparison of Pyridine- and PPh₂-Chelated Complexes 8
1.8. PPh₂-Chelated ImPy Ni(II) Complexes for Acrylate Synthesis 10
1.9. N2-Position for Steric Tuning 12
1.10. Ligand Designs with an Unsymmetric Steric Environments 14
1.10.1. F-ImPy-Ru Catalyst with Unsymmetric N-Aryl Ring 15
1.10.2. Ipaul Ligand with Unsymmetric and Rotatable N-Aryl Ring 16
1.10.3. Bulky yet Flexible Ligand Design 17
Research Purpose 19
Results and Discussion 23
3.1. Synthesis of Ligand Precursors 23
3.2. X-ray crysrallography 24
3.3. Ni(II) Complexes Bearing Unsymmetric PPh₂–ImPy Ligands
- Steric Map & Buried Volume Analysis 26
3.4. Catalytic Performances in Acrylate Synthesis 27
3.5. Potential Steric Effects of Unsymmetric N-Aryl Ring in the Acrylate Synthesis Catalysis. 29
Conclusion 31
Structure refinement data 32
Experimental Section 33
6.1. General Information 33
6.2. Synthesis of Unsymmetric Anilines 33
6.3. Synthesis of Imines with an Unsymmetric N-Aryl Ring 35
6.4. Synthesis of Phosphine-Chelated ImPy Ligand Precursors 38
6.5. Synthesis of Phosphine-Chelated ImPy Ni(II) Complex 39
6.6. Acrylate Synthesis from Ethylene and CO₂ 40
6.7. NMR Spectra 41
• References 47
• Acknowledgements 5
