Reconfigurable RCS Reduction from Curved Structures Using Plasma Based FSS

Plasmonics - The radar cross section (RCS) reduction from curved surfaces using plasma based frequency selective surfaces (FSS) is investigated. A frequency reconfigurable plasma based FSS...     

Thermal Transport Characteristics of Human Skin Measured In Vivo Using Ultrathin Conformal Arrays of Thermal Sensors and Actuators

Measurements of the thermal transport properties of the skin can reveal changes in physical and chemical states of relevance to dermatological health, skin structure and activity, thermoregulation and other aspects of human physiology. Existing methods for in vivo evaluations demand complex systems for laser heating and infrared thermography, or they require rigid, invasive probes; neither can apply to arbitrary regions of the body, offers modes for rapid spatial mapping, or enables continuous monitoring outside of laboratory settings. Here we describe human clinical studies using mechanically soft arrays of thermal actuators and sensors that laminate onto the skin to provide rapid, quantitative in vivo determination of both the thermal conductivity and thermal diffusivity, in a completely non-invasive manner. Comprehensive analysis of measurements on six different body locations of each of twenty-five human subjects reveal systematic variations and directional anisotropies in the characteristics, with correlations to the thicknesses of the epidermis (EP) and stratum corneum (SC) determined by optical coherence tomography, and to the water content assessed by electrical impedance based measurements. Multivariate statistical analysis establishes four distinct locations across the body that exhibit different physical properties: heel, cheek, palm, and wrist/volar forearm/dorsal forearm. The data also demonstrate that thermal transport correlates negatively with SC and EP thickness and positively with water content, with a strength of correlation that varies from region to region, e.g., stronger in the palmar than in the follicular regions.     

The Mechanism of Drag Reduction around Bodies of Revolution Using Bionic Non-Smooth Surfaces

Bionic non-smooth surfaces （BNSS） can reduce drag. Much attention has been paid to the mechanism of shear stress reduction by riblets. The mechanism of pressure force reduction by bionic non-smooth surfaces on bodies of revolution has not been well investigated. In this work CFD simulation has revealed the mechanism of drag reduction by BNSS, which may work in three ways. First, BNSS on bodies of revolution may lower the surface velocity of the medium, which prevents the sudden speed up of air on the cross section. So the bottom pressure of the model would not be disturbed sharply, resulting in less energy loss and drag reduction. Second, the magnitude of vorticity induced by the bionic model becomes smaller because, due to the sculpturing, the growth of tiny air bubbles is avoided. Thus the large moment of inertia induced by large air bubble is reduced. The reduction of the vorticity could reduce the dissipation of the eddy. So the pressure force could also be reduced. Third, the thickness of the momentum layer on the model becomes less which, according to the relationship between the drag coefficient and the momentum thickness, reduces drag.     

Deciphering Enzyme Function Using Peptide Arrays

Enzymes are key molecules in signal-transduction pathways. However, only a small fraction of more than 500 human kinases, 300 human proteases and 200 human phosphatases is characterised so far. Peptide microarray based technologies for extremely efficient profiling of enzyme substrate specificity emerged in the last years. This technology reduces set-up time for HTS assays and allows the identification of downstream targets. Moreover, peptide microarrays enable optimisation of enzyme substrates. Focus of this review is on assay principles for measuring activities of kinases, phosphatases or proteases and on substrate identification/optimisation for kinases. Additionally, several examples for reliable identification of substrates for lysine methyl-transferases, histone deacetylases and SUMO-transferases are given. Finally, use of high-density peptide microarrays for the simultaneous profiling of kinase activities in complex biological samples like cell lysates or lysates of complete organisms is described. All published examples of peptide arrays used for enzyme profiling are summarised comprehensively.     

Object Placement Using Performance Surfaces

Heterogeneous parallel clusters of workstations are being used to solve many important computational problems. Scheduling parallel applications on the best collection of machines in a heterogeneous computing environment is a complex problem. Performance prediction is vital to good application performance in this environment since utilization of an ill-suited machine can slow the computation down significantly. This paper addresses the problem of network performance prediction. A new methodology for characterizing network links and application's need for network resources is developed which makes use of Performance Surfaces [3]. This Performance Surface abstraction is used to schedule a parallel application on resources where it will run most efficiently.     

Mapping Genomic Library Clones Using Oligonucleotide Arrays

We have developed a high-density DNA probe array and accompanying biochemical and informatic methods to order clones from genomic libraries. This approach involves a series of enzymatic steps for capturing a set of short dispersed sequence markers scattered throughout a high-molecular-weight DNA. By this process, all the ambiguous sequences lying adjacent to a given Type IIS restriction site are ligated between two DNA adapters. These markers, once amplified and labeled by PCR, can be hybridized and detected on a high-density oligonucleotide array bearing probes complementary to all possible markers. The array is synthesized using light-directed combinatorial chemistry. For each clone in a genomic library, a characteristic set of sequence markers can be determined. On the basis of the similarity between the marker sets for each pair of clones, their relative overlap can be measured. The library can be sequentially ordered into a contig map using this overlap information. This new methodology does not require gel-based methods or prior sequence information and involves manipulations that should allow for easy adaptation to automated processing and data collection.     

Identifying Protein-protein Interaction Sites Using Peptide Arrays

Protein-protein interactions mediate most of the processes in the living cell and control homeostasis of the organism. Impaired protein interactions may result in disease, making protein interactions important drug targets. It is thus highly important to understand these interactions at the molecular level. Protein interactions are studied using a variety of techniques ranging from cellular and biochemical assays to quantitative biophysical assays, and these may be performed either with full-length proteins, with protein domains or with peptides. Peptides serve as excellent tools to study protein interactions since peptides can be easily synthesized and allow the focusing on specific interaction sites. Peptide arrays enable the identification of the interaction sites between two proteins as well as screening for peptides that bind the target protein for therapeutic purposes. They also allow high throughput SAR studies. For identification of binding sites, a typical peptide array usually contains partly overlapping 10-20 residues peptides derived from the full sequences of one or more partner proteins of the desired target protein. Screening the array for binding the target protein reveals the binding peptides, corresponding to the binding sites in the partner proteins, in an easy and fast method using only small amount of protein.In this article we describe a protocol for screening peptide arrays for mapping the interaction sites between a target protein and its partners. The peptide array is designed based on the sequences of the partner proteins taking into account their secondary structures. The arrays used in this protocol were Celluspots arrays prepared by INTAVIS Bioanalytical Instruments. The array is blocked to prevent unspecific binding and then incubated with the studied protein. Detection using an antibody reveals the binding peptides corresponding to the specific interaction sites between the proteins.     

Manipulation of Microscale Fluid Using Laser-Irradiated Nanoparticle Arrays

The manipulation of microscale fluid has been widely used in biology, medicine, and chemistry. However, the traditional control systems are relatively large, complex, and costly. Optical driving micro- and nanofluid is a new trend of microfluidics, which combines the advantages of both optics and microfluidics in the micro-nano scale. In the present work, we investigated a method to drive microfluid by taking advantage of the localized surface plasmon resonance effect of gold nanoparticles, which can convert optical energy to fluid motion. First, numerical simulation was carried out to calculate the electromagnetic, temperature, and flow field around laser-irradiated gold nanoparticles. Then, the simplified heat source condition was verified. The nanoparticle array was regarded as heat source to induce convection flow. The influence of the spacing and number of nanoparticles in array was investigated. On this basis, the structural parameters of nanoparticle array that can be used to regulate the velocity of microfluidic were obtained.

     

Using Affordable LED Arrays for Photo-Stimulation of Neurons

Standard slice electrophysiology has allowed researchers to probe individual components of neural circuitry by recording electrical responses of single cells in response to electrical or pharmacological manipulations^1,2. With the invention of methods to optically control genetically targeted neurons (optogenetics), researchers now have an unprecedented level of control over specific groups of neurons in the standard slice preparation. In particular, photosensitive channelrhodopsin-2 (ChR2) allows researchers to activate neurons with light^3,4. By combining careful calibration of LED-based photostimulation of ChR2 with standard slice electrophysiology, we are able to probe with greater detail the role of adult-born interneurons in the olfactory bulb, the first central relay of the olfactory system. Using viral expression of ChR2-YFP specifically in adult-born neurons, we can selectively control young adult-born neurons in a milieu of older and mature neurons. Our optical control uses a simple and inexpensive LED system, and we show how this system can be calibrated to understand how much light is needed to evoke spiking activity in single neurons. Hence, brief flashes of blue light can remotely control the firing pattern of ChR2-transduced newborn cells.Download video file.^{(48M, mov)}     

Predicting Antidisease Immunity Using Proteome Arrays and Sera from Children Naturally Exposed to Malaria

Malaria remains one of the most prevalent and lethal human infectious diseases worldwide. A comprehensive characterization of antibody responses to blood stage malaria is essential to support the development of future vaccines, sero-diagnostic tests, and sero-surveillance methods. We constructed a proteome array containing 4441 recombinant proteins expressed by the blood stages of the two most common human malaria parasites, P. falciparum (Pf) and P. vivax (Pv), and used this array to screen sera of Papua New Guinea children infected with Pf, Pv, or both (Pf/Pv) that were either symptomatic (febrile), or asymptomatic but had parasitemia detectable via microscopy or PCR. We hypothesized that asymptomatic children would develop antigen-specific antibody profiles associated with antidisease immunity, as compared with symptomatic children. The sera from these children recognized hundreds of the arrayed recombinant Pf and Pv proteins. In general, responses in asymptomatic children were highest in those with high parasitemia, suggesting that antibody levels are associated with parasite burden. In contrast, symptomatic children carried fewer antibodies than asymptomatic children with infections detectable by microscopy, particularly in Pv and Pf/Pv groups, suggesting that antibody production may be impaired during symptomatic infections. We used machine-learning algorithms to investigate the relationship between antibody responses and symptoms, and we identified antibody responses to sets of Plasmodium proteins that could predict clinical status of the donors. Several of these antibody responses were identified by multiple comparisons, including those against members of the serine enriched repeat antigen family and merozoite protein 4. Interestingly, both P. falciparum serine enriched repeat antigen-5 and merozoite protein 4 have been previously investigated for use in vaccines. This machine learning approach, never previously applied to proteome arrays, can be used to generate a list of potential seroprotective and/or diagnostic antigens candidates that can be further evaluated in longitudinal studies.Of the five species of malaria parasites that infect humans, Plasmodium falciparum (Pf)¹ and P. vivax (Pv) are the most common. Interventions aimed at reducing transmission and improving diagnosis and treatment have led to a dramatic reduction in morbidity and mortality (1, 2). For example, Pf fatalities have declined from an estimated one million to 655,000 annually (2). Although Pv is now recognized as the most widespread species worldwide and a significant cause of severe disease, this parasite, which can relapse months to years after the initial blood stage infection, is still largely ignored (3, 4). Furthermore, mixed-species infections, most commonly of Pf and Pv, are more frequent than previously thought. Although blood smears suggest that <2% of cases are mixed-species infections, PCR-based diagnoses suggest that 55–65% of infections in Thailand, Papua New Guinea (PNG), and other countries in south-east Asia (5–7) are mixed-species infections.Natural immunity can be subdivided into antidisease immunity and antiparasitic immunity. Antidisease immunity (defined as the absence of symptoms) develops quickly, sometimes requiring only one or two infections in high transmission areas (8–11). However, individuals living in high transmission areas develop non-sterile antiparasite immunity, resulting in low-level parasitemia and asymptomatic infections. This immunity is acquired much more slowly than antidisease immunity, may require repeated infections depending on the transmission rate, and is rarely sterilizing (12). Parasite densities in individuals that have acquired antiparasite immunity are average 10⁴- to 10⁶-fold lower than those in non-immune individuals (13).Blood stage parasites activate innate responses, which in turn lead to significant levels of humoral and cellular adaptive immunity (reviewed in (14)). Antiparasitic immunity appears to be mediated primarily by antibody responses against blood stage antigens (15, 16). Specific antimalarial antibodies can block invasion of host erythrocytes in vitro by both Pf (17) and Pv merozoites (18–21). Additionally, certain antibody isotypes, in particular IgG3, can induce antibody-dependent cellular inhibition (ADCI) of parasite invasion and development in erythrocytes, which is strongly associated with protection against malaria parasites (13, 22). Moreover, passive transfer of Pf antimalarial antibodies to infected patients can result in parasite clearance (15, 16). Evidence from field studies suggests that the slow acquisition of antibodies to genetically variant circulating strains over several years is associated with antidisease immunity to Pf (23), but to a lesser extent to Pv (24). Cell-mediated immune responses also play a role in protection, particularly early in the immune response. A strong pro-inflammatory response mediated primarily by interferon-gamma (IFN-γ) and tumor necrosis factor-α (TNF-α) contributes to the initial killing and clearance of parasite-infected red blood cells (25).Identifying antibody targets that are associated with infection, disease, or immunity will support the development of vaccines, diagnostics, and tools for sero-surveillance. By comparing the humoral response profiles of defined populations possessing varying degrees of antidisease and/or antiparasite immunity, it may be possible to identify combinations or responses that are associated with protection against clinical disease and/or parasitemia. These responses could guide selection of antigen(s) for blood stage vaccines. Here, we applied Plasmodium genome sequence, proteomics, bioinformatics, and proteome array fabrication technologies to construct a Pf/Pv blood stage proteome array. The Plasmodium genome encodes over 5000 proteins (5538 and 5435 in Pf and Pv, respectively (26)), nearly half of which have been identified via proteomics at the different stages of malaria parasite life cycle (27, 28). A total of 4441 recombinant proteins, representing 1922 Pf and 1936 Pv native proteins previously reported or predicted to be expressed by the blood stages of these parasites were included on the Pf/Pv proteome arrays, which were then used to analyze antibody responses to both Plasmodium species in naturally-exposed individuals with clinically characterized infections.The resulting data were interrogated using machine-learning algorithms to identify antibody responses that associated with disease status. We identified sets of antigen-specific antibody responses that can be used to distinguish between asymptomatic donors with parasitemias detectable by light microscopy or PCR and asymptomatic donors, some of which were identified by multiple comparisons. This study is a proof-of-concept of the power of applying machine learning algorithms to biomarker discovery, and paves the way for future, more robust studies to identify novel malaria vaccine targets.     

A 2-Bit Pancharatnam-Berry Coding Metasurface for Ultra-wideband and Polarization Insensitive RCS Reduction

Plasmonics - Because Pancharatnam-Berry (PB) geometrical phase can only be generated in the co-polarized reflection coefficient under circular polarized (CP) incidence for a reflective metasurface,...     

Stabilizing Hepatocellular Phenotype Using Optimized Synthetic Surfaces

Currently, one of the major limitations in cell biology is maintaining differentiated cell phenotype. Biological matrices are commonly used for culturing and maintaining primary and pluripotent stem cell derived hepatocytes. While biological matrices are useful, they permit short term culture of hepatocytes, limiting their widespread application. We have attempted to overcome the limitations using a synthetic polymer coating. Polymers represent one of the broadest classes of biomaterials and possess a wide range of mechanical, physical and chemical properties, which can be fine-tuned for purpose. Importantly, such materials can be scaled to quality assured standards and display batch-to-batch consistency. This is essential if cells are to be expanded for high through-put screening in the pharmaceutical testing industry or for cellular based therapy. Polyurethanes (PUs) are one group of materials that have shown promise in cell culture. Our recent progress in optimizing a polyurethane coated surface, for long-term culture of human hepatocytes displaying stable phenotype, is presented and discussed.     

Copper Reduction and Contact Killing of Bacteria by Iron Surfaces

The well-established killing of bacteria by copper surfaces, also called contact killing, is currently believed to be a combined effect of bacterial contact with the copper surface and the dissolution of copper, resulting in lethal bacterial damage. Iron can similarly be released in ionic form from iron surfaces and would thus be expected to also exhibit contact killing, although essentially no contact killing is observed by iron surfaces. However, we show here that the exposure of bacteria to iron surfaces in the presence of copper ions results in efficient contact killing. The process involves reduction of Cu²⁺ to Cu⁺ by iron; Cu⁺ has been shown to be considerably more toxic to cells than Cu²⁺. The specific Cu⁺ chelator, bicinchoninic acid, suppresses contact killing by chelating the Cu⁺ ions. These findings underline the importance of Cu⁺ ions in the contact killing process and infer that iron-based alloys containing copper could provide novel antimicrobial materials.     

Vascular Gene Transfer from Metallic Stent Surfaces Using Adenoviral Vectors Tethered through Hydrolysable Cross-linkers

In-stent restenosis presents a major complication of stent-based revascularization procedures widely used to re-establish blood flow through critically narrowed segments of coronary and peripheral arteries. Endovascular stents capable of tunable release of genes with anti-restenotic activity may present an alternative strategy to presently used drug-eluting stents. In order to attain clinical translation, gene-eluting stents must exhibit predictable kinetics of stent-immobilized gene vector release and site-specific transduction of vasculature, while avoiding an excessive inflammatory response typically associated with the polymer coatings used for physical entrapment of the vector. This paper describes a detailed methodology for coatless tethering of adenoviral gene vectors to stents based on a reversible binding of the adenoviral particles to polyallylamine bisphosphonate (PABT)-modified stainless steel surface via hydrolysable cross-linkers (HC). A family of bifunctional (amine- and thiol-reactive) HC with an average t_1/2 of the in-chain ester hydrolysis ranging between 5 and 50 days were used to link the vector with the stent. The vector immobilization procedure is typically carried out within 9 hr and consists of several steps: 1) incubation of the metal samples in an aqueous solution of PABT (4 hr); 2) deprotection of thiol groups installed in PABT with tris(2-carboxyethyl) phosphine (20 min); 3) expansion of thiol reactive capacity of the metal surface by reacting the samples with polyethyleneimine derivatized with pyridyldithio (PDT) groups (2 hr); 4) conversion of PDT groups to thiols with dithiothreitol (10 min); 5) modification of adenoviruses with HC (1 hr); 6) purification of modified adenoviral particles by size-exclusion column chromatography (15 min) and 7) immobilization of thiol-reactive adenoviral particles on the thiolated steel surface (1 hr). This technique has wide potential applicability beyond stents, by facilitating surface engineering of bioprosthetic devices to enhance their biocompatibility through the substrate-mediated gene delivery to the cells interfacing the implanted foreign material.     

Proteome-wide Epitope Mapping of Antibodies Using Ultra-dense Peptide Arrays

Antibodies are of importance for the field of proteomics, both as reagents for imaging cells, tissues, and organs and as capturing agents for affinity enrichment in mass-spectrometry-based techniques. It is important to gain basic insights regarding the binding sites (epitopes) of antibodies and potential cross-reactivity to nontarget proteins. Knowledge about an antibody''s linear epitopes is also useful in, for instance, developing assays involving the capture of peptides obtained from trypsin cleavage of samples prior to mass spectrometry analysis. Here, we describe, for the first time, the design and use of peptide arrays covering all human proteins for the analysis of antibody specificity, based on parallel in situ photolithic synthesis of a total of 2.1 million overlapping peptides. This has allowed analysis of on- and off-target binding of both monoclonal and polyclonal antibodies, complemented with precise mapping of epitopes based on full amino acid substitution scans. The analysis suggests that linear epitopes are relatively short, confined to five to seven residues, resulting in apparent off-target binding to peptides corresponding to a large number of unrelated human proteins. However, subsequent analysis using recombinant proteins suggests that these linear epitopes have a strict conformational component, thus giving us new insights regarding how antibodies bind to their antigens.Antibodies are used in proteomics both as imaging reagents for the analysis of tissue specificity (1) and subcellular localization (2) and as capturing agents for targeted proteomics (3), in particular for the enrichment of peptides for immunoaffinity methods such as Stable Isotope Standards and Capture by Anti-peptide Antibodies (4). In fact, the Human Proteome Project (5) has announced that one of the three pillars of the project will be antibody-based, with one of the aims being to generate antibodies to at least one representative protein from all protein-coding genes. Knowledge about the binding site (epitope) of an antibody toward a target protein is thus important for gaining basic insights into antibody specificity and sensitivity and facilitating the identification and design of antigens to be used for reagents in proteomics, as well as for the generation of therapeutic antibodies and vaccines (1, 6). With over 20 monoclonal-antibody-based drugs now on the market and over 100 in clinical trials, the field of antibody therapeutics has become a central component of the pharmaceutical industry (7). One of the key parameters for antibodies includes the nature of the binding recognition toward the target, involving either linear epitopes formed by consecutive amino acid residues or conformational epitopes consisting of amino acids brought together by the fold of the target protein (8).A large number of methods have therefore been developed to determine the epitopes of antibodies, including mass spectrometry (9), solid phase libraries (10, 11), and different display systems (12–14) such as bacterial display (15) and phage display (16). The most common method for epitope mapping involves the use of soluble and immobilized (tethered) peptide libraries, often in an array format, exemplified by the “Geysen Pepscan” method (11) in which overlapping “tiled” peptides are synthesized and used for binding analysis. The tiled peptide approach can also be combined with alanine scans (17) in which alanine substitutions are introduced into the synthetic peptides and the direct contribution of each amino acid can be investigated. Maier et al. (18) described a high-throughput epitope-mapping screen of a recombinant peptide library consisting of a total of 2304 overlapping peptides of the vitamin D receptor, and recently Buus et al. (19) used in situ synthesis on microarrays to design and generate 70,000 peptides for epitope mapping of antibodies using a range of peptides with sizes from 4-mer to 20-mer.So far it has not been possible to investigate on- and off-target binding in a proteome-wide manner, but the emergence of new methods for in situ synthesis of peptides on ultra-dense arrays has made this achievable. Here, we describe the design and use of peptide arrays generated with parallel in situ photolithic synthesis (20) of a total of 2.1 million overlapping peptides covering all human proteins with overlapping peptides. Miniaturization of the peptide arrays (21) has led to improved density of the synthesized peptides and consequently has improved the resolution and coverage of the epitope mapping. This has allowed us to study the specificity and cross-reactivity of both monoclonal and polyclonal antibodies across the whole “epitome” with the use of both proteome-wide arrays and focused-content peptide arrays covering selected antigen sequences to precisely map the contribution of each amino acid of the target protein for binding recognition of the corresponding antibodies. The results show the usefulness of proteome-wide epitope mapping, showing a path forward for high-throughput analysis of antibody interactions.     

Electrochemical CO2 Reduction Using Electrons Generated from Photoelectrocatalytic Phenol Oxidation

Electrochemical CO₂ reduction (ECCO₂R) requires electrons, for example, from oxygen evolution reaction (OER). However, such a multiple‐electron‐involved reaction is complicated and kinetically slow, leading to high overpotential. Herein, OER is replaced with photoelectrocatalytic phenol oxidation reaction (PECPOR) that provides electrons for ECCO₂R. In an integrated cell, ECCO₂R is conducted on the cathode of Cu nanowires and PECPOR is performed on the anode of SnO₂ and Sb coated TiO₂ nanotubes. Significant improvement of ECCO₂R into CO and hydrocarbons is realized when PECPOR is conducted at a high current density. The use of this integrated system results in the reduction of the specific energy consumption by a factor of 51.33%, compared with the utilization of two half‐cells for individual ECCO₂R and PECPOR. This study thus proposes a novel strategy to couple ECCO₂R with PECPOR and eventually to tackle the problems of environmental pollution and energy crisis.     

Optical Fiber-Based Surface-Enhanced Raman Scattering Sensor Using Au Nanovoid Arrays

Plasmonic properties of gold nanovoid array substrates for fiber-based surface-enhanced Raman scattering (SERS) sensing are studied numerically and experimentally. In the nanovoid arrays, each void has openings on both sides, bottom hole facing the fiber tip for introducing incident light and collecting scattered light and the top hole exposed to the analyte solution for interrogating analyte molecules in the voids. Electromagnetic field modes are confined strongly in and around these nanovoids, acting as localized plasmon resonators. The enhanced electric field extends hundreds of nanometers into the voids, resulting in a large SERS-active zone several orders of magnitude larger than nanoparticle-based structures. The effect of structural parameters of the nanovoid arrays, including void diameter, Au film thickness, and bottom hole diameter, on electric field confinement in the voids is investigated using three-dimensional finite difference time domain simulation. Au nanovoid arrays are fabricated using a scalable, inexpensive nanosphere lithography fabrication method. The largest SERS signal is realized by tuning the localized plasmon resonance peak of Au nanovoid arrays to the laser excitation wavelength. Multiplexed detection capability with the fiber-based SERS sensor using Au nanovoid arrays is demonstrated by measuring the Raman spectrum of a mixture solution of diethylthiatricarbocyanine and crystal violet.     

Specificity Analysis of Protein Lysine Methyltransferases Using SPOT Peptide Arrays

Lysine methylation is an emerging post-translation modification and it has been identified on several histone and non-histone proteins, where it plays crucial roles in cell development and many diseases. Approximately 5,000 lysine methylation sites were identified on different proteins, which are set by few dozens of protein lysine methyltransferases. This suggests that each PKMT methylates multiple proteins, however till now only one or two substrates have been identified for several of these enzymes. To approach this problem, we have introduced peptide array based substrate specificity analyses of PKMTs. Peptide arrays are powerful tools to characterize the specificity of PKMTs because methylation of several substrates with different sequences can be tested on one array. We synthesized peptide arrays on cellulose membrane using an Intavis SPOT synthesizer and analyzed the specificity of various PKMTs. Based on the results, for several of these enzymes, novel substrates could be identified. For example, for NSD1 by employing peptide arrays, we showed that it methylates K44 of H4 instead of the reported H4K20 and in addition H1.5K168 is the highly preferred substrate over the previously known H3K36. Hence, peptide arrays are powerful tools to biochemically characterize the PKMTs.     

Analysis of East Asia Genetic Substructure Using Genome-Wide SNP Arrays

Accounting for population genetic substructure is important in reducing type 1 errors in genetic studies of complex disease. As efforts to understand complex genetic disease are expanded to different continental populations the understanding of genetic substructure within these continents will be useful in design and execution of association tests. In this study, population differentiation (Fst) and Principal Components Analyses (PCA) are examined using >200 K genotypes from multiple populations of East Asian ancestry. The population groups included those from the Human Genome Diversity Panel [Cambodian, Yi, Daur, Mongolian, Lahu, Dai, Hezhen, Miaozu, Naxi, Oroqen, She, Tu, Tujia, Naxi, Xibo, and Yakut], HapMap [ Han Chinese (CHB) and Japanese (JPT)], and East Asian or East Asian American subjects of Vietnamese, Korean, Filipino and Chinese ancestry. Paired Fst (Wei and Cockerham) showed close relationships between CHB and several large East Asian population groups (CHB/Korean, 0.0019; CHB/JPT, 00651; CHB/Vietnamese, 0.0065) with larger separation with Filipino (CHB/Filipino, 0.014). Low levels of differentiation were also observed between Dai and Vietnamese (0.0045) and between Vietnamese and Cambodian (0.0062). Similarly, small Fst''s were observed among different presumed Han Chinese populations originating in different regions of mainland of China and Taiwan (Fst''s <0.0025 with CHB). For PCA, the first two PC''s showed a pattern of relationships that closely followed the geographic distribution of the different East Asian populations. PCA showed substructure both between different East Asian groups and within the Han Chinese population. These studies have also identified a subset of East Asian substructure ancestry informative markers (EASTASAIMS) that may be useful for future complex genetic disease association studies in reducing type 1 errors and in identifying homogeneous groups that may increase the power of such studies.