Browse
Submit Data
Databases
API
Help

Dataset Information

15 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

DNA-mediated assembly of protein heterodimers on membrane surfaces.

ABSTRACT: We present a method based on self-assembling oligonucleotides to anchor proteins to a supported membrane surface. This anchoring method allows control of the surface density of multiple proteins. By incorporating additional recognition sequences into the DNA linkers, defined heterodimers can be produced upon the addition of a heterospecific DNA cross-linking strand. Characterization by fluorescence cross-correlation spectroscopy (FCCS) confirmed lateral mobility and the formation of specific heterodimers. We further demonstrate that proteins linked in this manner as either monomers or dimers can form functional interfaces with living cells.

SUBMITTER: Coyle MP

PROVIDER: S-EPMC3626235 | biostudies-literature | 2013 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

DNA-mediated assembly of protein heterodimers on membrane surfaces.

Coyle Michael P MP Xu Qian Q Chiang Samantha S Francis Matthew B MB Groves Jay T JT

Journal of the American Chemical Society 20130326 13

We present a method based on self-assembling oligonucleotides to anchor proteins to a supported membrane surface. This anchoring method allows control of the surface density of multiple proteins. By incorporating additional recognition sequences into the DNA linkers, defined heterodimers can be produced upon the addition of a heterospecific DNA cross-linking strand. Characterization by fluorescence cross-correlation spectroscopy (FCCS) confirmed lateral mobility and the formation of specific het ...[more]

PMID: 23530555

Similar Datasets

Kinetic analysis of cytokine-mediated receptor assembly using engineered FC heterodimers.

Project description:A method for analyzing ligand-receptor binding kinetics is described, which is based on an engineered FC domain (FChk) that forms a covalent heterodimer. To validate the system, the type I IFN receptors (IFNAR1 and IFNAR2) were expressed as IFNAR1-FChk, IFNAR2-FCkh, and IFNAR1/IFNAR2-FChk fusion proteins. Surface plasmon resonance (SPR) analysis of binary IFN?2a/IFNAR interactions confirmed prior affinity measurements, while the affinity of the IFN?2a/IFNAR1/IFNAR2-FChk interaction reproduced the affinity of IFN?2a binding to living cells. In cellular assays, IFNAR1/IFNAR2-FChk potently neutralized IFN?2a bioactivity with an inhibitory concentration equivalent to the KD measured by SPR. These studies suggest that FChk provides a simple reagent to evaluate the binding kinetics of multiple ligand-receptor signaling systems that control cell growth, development, and immunity.

| S-EPMC3832046 | biostudies-literature

Direct membrane protein-DNA interactions required early in nuclear envelope assembly.

Project description:Among the earliest events in postmitotic nuclear envelope (NE) assembly are the interactions between chromatin and the membranes that will fuse to form the NE. It has been proposed that interactions between integral NE proteins and chromatin proteins mediate initial membrane recruitment to chromatin. We show that several transmembrane NE proteins bind to DNA directly and that NE membrane proteins as a class are enriched in long, basic domains that potentially bind DNA. Membrane fractions that are essential for NE formation are shown to bind directly to protein-free DNA, and our data suggest that these interactions are critical for early steps in NE assembly.

| S-EPMC2063857 | biostudies-literature

Morphological changes of plasma membrane and protein assembly during clathrin-mediated endocytosis.

Project description:Clathrin-mediated endocytosis (CME) proceeds through a series of morphological changes of the plasma membrane induced by a number of protein components. Although the spatiotemporal assembly of these proteins has been elucidated by fluorescence-based techniques, the protein-induced morphological changes of the plasma membrane have not been fully clarified in living cells. Here, we visualize membrane morphology together with protein localizations during CME by utilizing high-speed atomic force microscopy (HS-AFM) combined with a confocal laser scanning unit. The plasma membrane starts to invaginate approximately 30 s after clathrin starts to assemble, and the aperture diameter increases as clathrin accumulates. Actin rapidly accumulates around the pit and induces a small membrane swelling, which, within 30 s, rapidly covers the pit irreversibly. Inhibition of actin turnover abolishes the swelling and induces a reversible open-close motion of the pit, indicating that actin dynamics are necessary for efficient and irreversible pit closure at the end of CME.

| S-EPMC5953504 | biostudies-literature

DNA and protein isolation from macrophyte surfaces

Project description:Selective DNA and protein isolation from marine macrophyte surfaces

| PRJEB37845 | ENA

Protein-Mediated Colloidal Assembly.

Project description:Programmable colloidal assembly enables the creation of mesoscale materials in a bottom-up manner. Although DNA oligonucleotides have been used extensively as the programmable units in this paradigm, proteins, which exhibit more diverse modes of association and function, have not been widely used to direct colloidal assembly. Here we use protein-protein interactions to drive controlled aggregation of polystyrene microparticles, either through reversible coiled-coil interactions or through intermolecular isopeptide linkages. The sizes of the resulting aggregates are tunable and can be controlled by the concentration of immobilized surface proteins. Moreover, particles coated with different protein pairs undergo orthogonal assembly. We demonstrate that aggregates formed by association of coiled-coil proteins, in contrast to those linked by isopeptide bonds, are dispersed by treatment with chemical denaturants or soluble competing proteins. Finally, we show that protein-protein interactions can be used to assemble complex core-shell aggregates. This work illustrates a versatile strategy for engineering colloidal systems for use in materials science and biotechnology.

| S-EPMC5672941 | biostudies-literature

H-Ras forms dimers on membrane surfaces via a protein-protein interface.

Project description:The lipid-anchored small GTPase Ras is an important signaling node in mammalian cells. A number of observations suggest that Ras is laterally organized within the cell membrane, and this may play a regulatory role in its activation. Lipid anchors composed of palmitoyl and farnesyl moieties in H-, N-, and K-Ras are widely suspected to be responsible for guiding protein organization in membranes. Here, we report that H-Ras forms a dimer on membrane surfaces through a protein-protein binding interface. A Y64A point mutation in the switch II region, known to prevent Son of sevenless and PI3K effector interactions, abolishes dimer formation. This suggests that the switch II region, near the nucleotide binding cleft, is either part of, or allosterically coupled to, the dimer interface. By tethering H-Ras to bilayers via a membrane-miscible lipid tail, we show that dimer formation is mediated by protein interactions and does not require lipid anchor clustering. We quantitatively characterize H-Ras dimerization in supported membranes using a combination of fluorescence correlation spectroscopy, photon counting histogram analysis, time-resolved fluorescence anisotropy, single-molecule tracking, and step photobleaching analysis. The 2D dimerization Kd is measured to be ∼1 × 10(3) molecules/µm(2), and no higher-order oligomers were observed. Dimerization only occurs on the membrane surface; H-Ras is strictly monomeric at comparable densities in solution. Analysis of a number of H-Ras constructs, including key changes to the lipidation pattern of the hypervariable region, suggest that dimerization is a general property of native H-Ras on membrane surfaces.

| S-EPMC3939930 | biostudies-literature

Altered Escherichia coli membrane protein assembly machinery allows proper membrane assembly of eukaryotic protein vitamin K epoxide reductase.

Project description:Functional overexpression of polytopic membrane proteins, particularly when in a foreign host, is often a challenging task. Factors that negatively affect such processes are poorly understood. Using the mammalian membrane protein vitamin K epoxide reductase (VKORc1) as a reporter, we describe a genetic selection approach allowing the isolation of Escherichia coli mutants capable of functionally expressing this blood-coagulation enzyme. The isolated mutants map to components of membrane protein assembly and quality control proteins YidC and HslV. We show that changes in the VKORc1 sequence and in the YidC hydrophilic groove along with the inactivation of HslV promote VKORc1 activity and dramatically increase its expression level. We hypothesize that such changes correct for mismatches in the membrane topogenic signals between E. coli and eukaryotic cells guiding proper membrane integration. Furthermore, the obtained mutants allow the study of VKORc1 reaction mechanisms, inhibition by warfarin, and the high-throughput screening for potential anticoagulants.

| S-EPMC4679017 | biostudies-literature

Subunit isoform selectivity in assembly of Na,K-ATPase ?-? heterodimers.

Project description:To catalyze ion transport, the Na,K-ATPase must contain one ? and one ? subunit. When expressed by transfection in various expression systems, each of the four ? subunit isoforms can assemble with each of the three ? subunit isoforms and form an active enzyme, suggesting the absence of selective ?-? isoform assembly. However, it is unknown whether in vivo conditions the ?-? assembly is random or isoform-specific. The ?(2)-?(2) complex was selectively immunoprecipitated by both anti-?(2) and anti-?(2) antibodies from extracts of mouse brain, which contains cells co-expressing multiple Na,K-ATPase isoforms. Neither ?(1)-?(2) nor ?(2)-?(1) complexes were detected in the immunoprecipitates. Furthermore, in MDCK cells co-expressing ?(1), ?(1), and ?(2) isoforms, a greater fraction of the ?(2) subunits was unassembled with ?(1) as compared with that of the ?(1) subunits, indicating preferential association of the ?(1) isoform with the ?(1) isoform. In addition, the ?(1)-?(2) complex was less resistant to various detergents than the ?(1)-?(1) complex isolated from MDCK cells or the ?(2)-?(2) complex isolated from mouse brain. Therefore, the diversity of the ?-? Na,K-ATPase heterodimers in vivo is determined not only by cell-specific co-expression of particular isoforms, but also by selective association of the ? and ? subunit isoforms.

| S-EPMC3406695 | biostudies-literature

Construction of Antifouling Membrane Surfaces through Layer-by-Layer Self-Assembly of Lignosulfonate and Polyethyleneimine.

Project description:Lignin is the second most abundant and low-cost natural polymer, but its high value-added utilization is still lack of effective and economic ways. In this paper, waste lignosulfonate (LS) was introduced to fabricate antifouling membrane surfaces via layer-by-layer self-assembly with polyethyleneimine (PEI). The LS/PEI multilayers were successfully deposited on the polysulfone (PSf) membrane, as demonstrated by ATR-FTIR, XPS, Zeta potential measurements, AFM, and SEM. Meanwhile, the effect of the number of bilayers was investigated in detail on the composition, morphologies, hydrophilicity, and antifouling performance of the membrane surface. As a result, with the bilayer numbers increase to 5, the PSf membrane shows smooth surface with small roughness, and its water contact angle reduces to 44.1°, indicating the improved hydrophilicity. Accordingly, the modified PSf membrane with 5 LS/PEI bilayers repels the adsorption of protein, resulting in good antifouling performance. This work provides a green, facile, and low-cost strategy to construct antifouling membrane surfaces.

| S-EPMC6918325 | biostudies-literature

Protein subunit interfaces: heterodimers versus homodimers.

Project description:Protein dimers are either homodimers (complexation of identical monomers) or heterodimers (complexation of non-identical monomers). These dimers are common in catalysis and regulation. However, the molecular principles of protein dimer interactions are difficult to understand mainly due to the geometrical and chemical characteristics of proteins. Nonetheless, the principles of protein dimer interactions are often studied using a dataset of 3D structural complexes determined by X-ray crystallography. A number of physical and chemical properties govern protein dimer interactions. Yet, a handful of such properties are known to dominate protein dimer interfaces. Here, we discuss the differences between homodimer and heterodimer interfaces using a selected set of interface properties.

| S-EPMC1891636 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data