Fasting plasma zeaxanthin response to Fructus barbarum L. (wolfberry; Kei Tze) in a food-based human supplementation trial

Antioxidant Research Group, Faculty of Health & Social Sciences2004-2005 > Academic research: refereed > Publication in refereed journalVersion of RecordPublishe

Seeing is believing; tracking metalloproteins by fluorescent probe in vivo and in vitro

Abstract no. EuAsC2S-12/S1-OP22Extensive genome research has shown that around 1/4 to 1/3 proteins are metalloproteins (or metal-binding proteins) with various metal ions incorporated with proteins for either structural or functional purposes. Thus, metalloproteomics/metallomics are developed to investigate the molecular mechanism of metal-related biological processes and the entirety of metal/metalloid species within a cell or tissue type[1]. Fluorescence labeling is probably the best method in view of its capability in providing rapid and sensitive identification in living biological systems. In spite of the development of fluorescent proteins, synthetic small-molecule fluorescence agents have been utilized to identify specific targets in cells, while metal-chelation methodology has been extensively applied to the study of metal-oriented biological process[2]. Although different types of metal-responsive sensors have been developed to label cellular metals[3], tracking of metal-binding proteins in living cells by fluorescence is still highly anticipated. In this work, novel fluorescent probe was designed to label metalloproteins both in vivo and in vitro. The protein partners of several metal ions such as Ni2+ (Histidine-rich proteins in particular), Bi3+, Cr3+ have been identified by the agent. The fluorescent agent exhibited “turnon” response to the targets in SDS-PAGE, and its excellent permeability enabled “lighting up” of targeted proteins in living cells, providing valuable information on metalloprotein spatial distribution in biology.postprintThe 12th EuroAsia Conference on Chemical Sciences (EuAsC2S-12), Corfu, Greece, 16-21 April 2012

Selective binding of Hpnl towards Ni(II) and Bi(III)

Poster-5Histidine-rich protein Hpn and histidine- and glutamine-rich protein Hpn-like (Hpnl) in Helicobacter pylori have been corroborated to be crucial to nickel homeostasis.[1-3] Nickel supply to hydrogenases and ureases might be disrupted owing to the interaction of metallodrugs, such as bismuth antiulcer drugs, with Hpnl, which may subsequently disturb the functions of the essential …postprin

An Intergrated Approach for Matching Metals and Metallodrugs to Proteins

Keynote Lecture (Abstract)The effect of metals in biology effects is double-edged. Metal ions operate, on one hand, as cofactors for around 40% enzymes, on the other hand, they also exhibit toxic effects. Some metal ions, although being not essential, have been widely used in human healthcare as either therapeutic agents or diagnosis agents. To understand the molecular mechanism of a metallodrug, it is crucial to match metals to proteins at a proteome-wide scale [1,2]. We used an integrated approach consisting of gel electrophoresis and inductively coupled plasma mass spectrometry, LA-ICP-MS, IMAC and bioinformatic approach to identify metal-associated proteins using bismuth antiulcer drug as an example [3,4]. Using continuous-flow gel electrophoresis in combination with ICP-MS, we developed a comprehensive and robust strategy to readily identify metal-associated proteins as well as to quantify the metals for fast metallome/proteome-wide profiling of metal-binding proteins. At the same time, we have developed a tunable fluorescent method to visualize metalbinding proteins and histidine-rich proteins directly in cells. To match metals to proteins, we also established a bioinformatic method which allows potential metal-binding proteins both sequentially and spaciously to be searched [5-7]. Surprisingly, histidine-rich proteins and motifs(HRMs) are commonly found in proteins. We systematically analyzed the proteomes of 675 prokaryotes including 50 archaea and 625 bacteria for HRMs, and show that HRMs are extensively distributed in prokaryotic proteomes, with the majority (62%) of histidine-rich proteins (HRPs) being involved in metal homeostasis. Importantly, the occurrence of histidine-rich proteins (motifs) in the proteomes of prokaryotes is related to their habitats.published_or_final_versio

EASY: efficient arbiter SYnthesis from multi-threaded code

High-Level Synthesis (HLS) tools automatically transform a high-level specification of a circuit into a low-level RTL description.Traditionally, HLS tools have operated on sequential code, howeverin recent years there has been a drive to synthesize multi-threadedcode. A major challenge facing HLS tools in this context is how toautomatically partition memory amongst parallel threads to fullyexploit the bandwidth available on an FPGA device and avoid mem-ory contention. Current automatic memory partitioning techniqueshave inefficient arbitration due to conservative assumptions regard-ing which threads may access a given memory bank. In this paper,we address this problem through formal verification techniques,permitting a less conservative, yet provably correct circuit to begenerated. We perform a static analysis on the code to determinewhich memory banks are shared by which threads. This analysisenables us to optimize the arbitration efficiency of the generatedcircuit. We apply our approach to the LegUp HLS tool and showthat for a set of typical application benchmarks we can achieve upto 87% area savings, and 39% execution time improvement, withlittle additional compilation time

Selective interaction of Hpn-like protein with nickel, zinc and bismuth in vitro and in cells by FRET

Hpn-like (Hpnl) is a unique histidine- and glutamine-rich protein found only in Helicobacter pylori and plays a role on nickel homeostasis.Weconstructed the fluorescent sensor proteins CYHpnl and CYHpnl_1-48 (C-terminal glutamine-rich region truncated) using enhanced cyan and yellow fluorescent proteins (eCFP and eYFP) as the donor–acceptor pair to monitor the interactions of Hpnl with metal ions and to elucidate the role of conserved Glu-rich sequence in Hpnl by fluorescence resonance energy transfer (FRET). CYHpnl and CYHpnl_1-48 exhibited largest responses towards Ni(II) and Zn(II) over other metals studied and the binding of Bi(III) to CYHpnl was observed in the presence of an excess amount of Bi(III) ions (Kd =115±4.8 μM). Moreover, both CYHpnl and CYHpnl_1-48 showed positive FRET responses towards the binding to Ni(II) and Zn(II) in Escherichia coli cells overexpressing CYHpnl and CYHpnl_1-48, whereas a decrease in FRET upon Bi(III)-binding in E. coli cells overexpressing the latter. Our study provides clear evidence on Hpnl binding to nickel in cells, and intracellular interaction of Hpnl with Bi(III) could disrupt the protein function, thus probably contributing to the efficacy of Bi(III) drugs against H. pylori.postprin

Efficient Memory Arbitration in High-Level Synthesis From Multi-Threaded Code

High-level synthesis (HLS) is an increasingly popular method for generating hardware from a description written in a software language like C/C++. Traditionally, HLS tools have operated on sequential code, however, in recent years there has been a drive to synthesise multi-threaded code. In this context, a major challenge facing HLS tools is how to automatically partition memory among parallel threads to fully exploit the bandwidth available on an FPGA device and minimise memory contention. Existingpartitioning approaches require inefficient arbitration circuitry to serialise accesses to each bank because they make conservative assumptions about which threads might access which memory banks. In this article, we design a static analysis that can prove certain memory banks are only accessed by certain threads, and use this analysis to simplify or even remove the arbiters while preserving correctness. We show how this analysis can be implemented using the Microsoft Boogie verifier on top of satisfiability modulo theories (SMT) solver, and propose a tool named EASY using automatic formal verification. Our work supports arbitrary input code with any irregular memory access patterns and indirect array addressing forms. We implement our approach in LLVM and integrate it into the LegUp HLS tool. For a set of typical application benchmarks our results have shown that EASY can achieve 0.13×(avg. 0.43×) of area and 1.64×(avg. 1.28×) of performance compared to the baseline, with little additional compilation time relative to the long time in hardware synthesis

The Diurnal Variation on Cardiovascular Endurance Performance of Secondary School Athlete Student

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Single Cell Transfection through Precise Microinjection with Quantitatively Controlled Injection Volumes

Cell transfection is a technique wherein foreign genetic molecules are delivered into cells. To elucidate distinct responses during cell genetic modification, methods to achieve transfection at the single-cell level are of great value. Herein, we developed an automated micropipette-based quantitative microinjection technology that can deliver precise amounts of materials into cells. The developed microinjection system achieved precise single-cell microinjection by pre-patterning cells in an array and controlling the amount of substance delivered based on injection pressure and time. The precision of the proposed injection technique was examined by comparing the fluorescence intensities of fluorescent dye droplets with a standard concentration and water droplets with a known injection amount of the dye in oil. Injection of synthetic modified mRNA (modRNA) encoding green fluorescence proteins or a cocktail of plasmids encoding green and red fluorescence proteins into human foreskin fibroblast cells demonstrated that the resulting green fluorescence intensity or green/red fluorescence intensity ratio were well correlated with the amount of genetic material injected into the cells. Single-cell transfection via the developed microinjection technique will be of particular use in cases where cell transfection is challenging and genetically modified of selected cells are desiredpublished_or_final_versio