Multifunctional Enzymes in Natural Product Biosynthesis

This cumulative doctoral thesis encompasses five research articles focused on investigations of multifunctional enzymes in natural product biosynthesis. The primary focus of this research is on exploring protein-protein and protein-substrate interactions, which play a critical role in polyketide and non-ribosomal peptide biosynthetic pathways. A part of this research is about studies of PPTase-CP interactions, ketosynthase-substrate, and dehydratase-substrate specificities. An interesting contribution of this thesis is the detailed investigation into the development of fluorescent probes in monitoring PPTase-CP interactions. This provides a simple way to monitor the carrier proteins activation by PPTases. Additionally, this thesis investigated the substrate specificity of ketosynthase by using 13C-labeled substrates. Another key enzyme explored in this thesis is dehydratase, which plays an important role in the polyketide and fatty acid biosynthetic pathways. In order to investigate the dehydratase product configurations and substrate scope, isotopically labeled probes and various substrates were synthesized, which enabled detailed stereochemical investigations. Moreover, this thesis also investigates the bifunctional terpene synthase, geosmin synthase, and presents the total synthesis of all isomers of geosmin, along with the characterization of the new compound, isogeosmin, which is produced by geosmin synthase. All in all, this thesis provides a comprehensive and detailed investigations of multifunctional enzymes in natural products biosynthesis. It advances the understanding of the enzymatic processes involved in these essential biochemical pathways, offering new insights into the enyzme mechanisms that drive these reactions

MICROSTRUCTURAL EVOLUTION AND MECHANICAL PROPERTIES OF Zn-Ti ALLOYS FOR BIODEGRADABLE STENT APPLICATIONS

Stents made of biodegradable metallic materials are increasingly gaining interest within the biomaterials field because of their superior mechanical properties and biodegradation rates as compared to polymeric materials. Zinc and its alloys have been developed and investigated as possible candidates for biodegradable stent applications in the last five years. This study intended to formulate and characterize a new series of Zn-Ti alloys, with titanium additions of less than 1-3 wt%, with the primary objective to develop and select an alloy that meets benchmark values of mechanical properties for biodegradable stents. A series of Zn-Ti alloys was formulated through vacuum induction melting. The experimental approach was to analyze the effect of Ti alloying element addition on mechanical properties of zinc. The structure, mechanical properties and fractography of the as-cast alloys were investigated. It was found that the grain size was reduced from above 600 µm to ~23 µm with the Ti content increasing from 0.01 wt% to 0.3 wt%. The amount of the intermetallic phase increased from 0.3 wt% to 2.5 wt% with Ti content. The results identify the formation of a eutectic phase of zinc with intermetallics at the primary grain boundaries. Zn16Ti was identified as the intermetallic phase formed in the as-cast Zn-Ti alloys. With increasing Ti content from 0.01 wt% to 1 wt%, the ultimate tensile strength and yield strength of the as cast Zn-Ti alloys increased from 101 and 64 MPa to 177 and 122 MPa, respectively. It is proposed that the strength of as-cast Zn-Ti alloys increases with the Ti content increasing from 0.01 wt% to 0.3 wt% due to grain refinement from a small percentage of titanium. The amount of the intermetallic phase increased with the Ti content increasing from 0.3 wt% to 2.5 wt%. It is proposed that the hardness and strength of the as-cast Zn-Ti alloys increased with the Ti content increasing from 0.3 to 2.5 wt% due to the increased formation of Zn-Ti intermetallic phases. The low elongation of the as-cast Zn-0.3 wt% Ti (3%), Zn-0.5 wt% Ti (4%), and Zn-1 wt% Ti alloys (2%) was also attributed to the increasing content of Zn-Ti intermetallic phases. Based on the results of the structure and mechanical properties of as-cast Zn-Ti alloys, the most promising as-cast candidates were processed through hot extrusion. This phase study was focused on the structure-property relationships before and after hot extrusion. The as-extruded Zn-0.01 wt% Ti had the highest average ultimate tensile strength and yield strength of 269 and 177 MPa, respectively. It is proposed that a significant increase in the ultimate tensile strength and yield strength in Zn-0.01 wt% Ti alloy after hot extrusion is due to grain refinement and formation of precipitates. The as-extruded Zn-0.1 wt% Ti and Zn-0.3 wt% Ti alloys exhibited high ductility, with the elongation to failure of about 44% and 30%, respectively. It is proposed that the as-extruded Zn-0.1 wt% Ti alloy exhibited high ductility due to the grain refinement and grain shape adjustment after hot extrusion. The high elongation of the as-extruded Zn-0.1 wt% Ti and Zn-0.3 wt% Ti alloys is consistent with the microstructural observations of ductile fracture. The as-extruded Zn-0.1 wt% Ti alloy had the best combination of tensile mechanical properties (UTS=207 MPa, YS=163 MPa, and Elongation=44%), which nearly meet the mechanical requirements for stent application

CD133: a potential indicator for differentiation and prognosis of human cholangiocarcinoma.

RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are.BACKGROUND: CD133 is known to be a cancer stem cell (CSC) marker. However, recent studies have revealed that CD133 is not restricted to CSC but to be expressed not only in human normal tissues but also in some cancers and could serve as a prognostic factor for the patients. Nevertheless, the expression of CD133 in human cholangiocarcinoma (CC) is rare and our study is to detect the expression and explore the potential functions of CD133 in human CC. METHODS: Fifty-nine cases, comprised of 5 normal liver tissues and 54 consecutive CC specimens (21 well-differentiated, 12 moderately-differentiated and 21 poorly-differentiated), were included in the study. Immunohistochemical stainning with CD133 protein was carried out, and statistical analyses were performed. RESULTS: CD133 was found to express in all 5 normal livers and 40 out of 54 (74%) CC tissues with different subcellular localization. In the well, moderately and poorly differentiated cases, the numbers of CD133 positive cases were 19 (19 of 21, 90%), 10 (10 of 12, 83%) and 11 (11 of 21, 52%) respectively. Further statistical analyses indicated that the expression and different subcellular localization of CD133 were significantly correlated with the differentiation status of tumors (P = 0.004, P = 0.009). Among 23 patients followed up for survival, the median survival was 4 months for fourteen CD133 negative patients but 14 months for nine CD133 positive ones. In univariate survival analysis, CD133 negative expression correlated with poor prognosis while CD133 positive expression predicted a favorable outcome of CC patients (P = 0.001). CONCLUSIONS: Our study demonstrates that CD133 expression correlates with the differentiation of CC and indicates that CD133 is a potential indicator for differentiation and prognosis of human CC.Published versio

Knowledge Distillation and Training Balance for Heterogeneous Decentralized Multi-Modal Learning over Wireless Networks

Decentralized learning is widely employed for collaboratively training models using distributed data over wireless networks. Existing decentralized learning methods primarily focus on training single-modal networks. For the decentralized multi-modal learning (DMML), the modality heterogeneity and the non-independent and non-identically distributed (non-IID) data across devices make it difficult for the training model to capture the correlated features across different modalities. Moreover, modality competition can result in training imbalance among different modalities, which can significantly impact the performance of DMML. To improve the training performance in the presence of non-IID data and modality heterogeneity, we propose a novel DMML with knowledge distillation (DMML-KD) framework, which decomposes the extracted feature into the modality-common and the modality-specific components. In the proposed DMML-KD, a generator is applied to learn the global conditional distribution of the modality-common features, thereby guiding the modality-common features of different devices towards the same distribution. Meanwhile, we propose to decrease the number of local iterations for the modalities with fast training speed in DMML-KD to address the imbalanced training. We design a balance metric based on the parameter variation to evaluate the training speed of different modalities in DMML-KD. Using this metric, we optimize the number of local iterations for different modalities on each device under the constraint of remaining energy on devices. Experimental results demonstrate that the proposed DMML-KD with training balance can effectively improve the training performance of DMML.Comment: submitted to IEEE Trans. on Mobile Computin

Aggregation Design for Personalized Federated Multi-Modal Learning over Wireless Networks

Federated Multi-Modal Learning (FMML) is an emerging field that integrates information from different modalities in federated learning to improve the learning performance. In this letter, we develop a parameter scheduling scheme to improve personalized performance and communication efficiency in personalized FMML, considering the non-independent and nonidentically distributed (non-IID) data along with the modality heterogeneity. Specifically, a learning-based approach is utilized to obtain the aggregation coefficients for parameters of different modalities on distinct devices. Based on the aggregation coefficients and channel state, a subset of parameters is scheduled to be uploaded to a server for each modality. Experimental results show that the proposed algorithm can effectively improve the personalized performance of FMML.Comment: accepted by IEEE Communications Letter

VACUUM BRAZING OF DIAMOND TO TUNGSTEN CARBIDE

Diamond tools are increasingly gaining importance as cutting and drilling materials for a wide variety of industrial applications. Polycrystalline diamond (PCD) is the main ultrahard material commercially used in the oil and gas drilling industry. In this study, a reactive brazing process was developed to join polycrystalline diamond (PCD) to WC-13 wt% Co, to form the cutter for fixed-cutter drill bit applications. Most nonmetals including polycrystalline diamond are not wet by and cannot easily be joined with conventional brazing alloys due to their chemical stability. The experimental approach was first to analyze the effect of adding an active metal (Ti, Zr, or V) to copper, silver, or a silver-copper eutectic alloy on the wettability of diamond and WC-Co substrates. Sessile drop tests were utilized to compare wettability between the liquid braze alloy and the substrate. The addition of Ti, Zr, and V decreased the apparent contact angle, which improved both the wetting and bonding behavior between braze alloy and diamond substrate. For all three alloy systems evaluated, all three base alloys (Cu, Ag, and Ag-Cu) with active metal additions (Ti, Zr, or V) exhibited good wettability on diamond and WC-Co substrates. Microstructural analysis of the diamond and WC-Co sessile drop samples was performed via scanning electron microscopy (SEM) to characterize the interfacial layers formed. Two different types of reactions were observed between the braze alloys and the WC-Co substrates: reduction and dissolution reactions. For the diamond sessile drop samples, only intermetallic solidification products were observed at the interface for the Ag-Cu eutectic based alloys with additions of 2 and 5 wt% Ti. SEM/EDS analysis revealed that the chemical changes at the interface between the braze alloy and diamond substrate were in agreement with the intermetallic solidification products predicted from the phase diagrams. Based on the Gibbs energies of formation for carbides, it is predicted that the formation of TiC is thermodynamically favored at the interface. However, no TiC reaction product was identified within the resolution of SEM/EDS analysis possibly because the TiC reaction layer is too thin. Based on the results of the wetting studies, an effort was made to optimize the shear strength of diamond brazed to WC-Co. This phase study was focused on the relationship between the braze alloy composition, the braze layer thickness, the brazing thermal cycle, the braze microstructures and the resulting joint mechanical properties. The average shear strength for Ag-2 wt% Ti alloy was approximately constant in the braze thickness range of 0.1 to 0.2 mm. It was observed that the brazed samples failed in the silver braze layer. More visible cracking and larger cracks were observed on the surface region of diamond substrates of the joint thickness of 0.2 mm for the Ag-Cu-2 wt% Ti alloys. It is possible that thermal stresses generated from coefficient of thermal expansion (CTE) mismatch resulted in the formation of interfacial cracks. The Ag-Cu eutectic alloy with addition of a 2 wt% Ti has the highest average shear strength of 95 MPa when the hold time is 30 minutes and the cooling rate is 5 °C/min