scholarly journals Electrodes modified with lipid membranes to study quinone oxidoreductases

2009 ◽  
Vol 37 (4) ◽  
pp. 707-712 ◽  
Author(s):  
Sophie A. Weiss ◽  
Lars J.C. Jeuken
Keyword(s):  
Lipid Membranes ◽  
Biochemical Properties ◽  
Model Membranes ◽  
Water Soluble ◽  
Enzyme Assays ◽  
Membrane Enzymes ◽  
Membrane Bound ◽  
Hydrophobic Nature

Quinone oxidoreductases are a class of membrane enzymes that catalyse the oxidation or reduction of membrane-bound quinols/quinones. The conversion of quinone/quinol by these enzymes is difficult to study because of the hydrophobic nature of the enzymes and their substrates. We describe some biochemical properties of quinones and quinone oxidoreductases and then look in more detail at two model membranes that can be used to study quinone oxidoreductases in a native-like membrane environment with their native lipophilic quinone substrates. The results obtained with these model membranes are compared with classical enzyme assays that use water-soluble quinone analogues.

scholarly journals Incomplete pneumolysin oligomers form membrane pores

Open Biology ◽  
2014 ◽  
Vol 4 (4) ◽  
pp. 140044 ◽  
Author(s):  
Andreas F.-P. Sonnen ◽  
Jürgen M. Plitzko ◽  
Robert J. C. Gilbert
Keyword(s):  
Lipid Membranes ◽  
Electron Tomography ◽  
Electrical Conductance ◽  
Membrane Attack Complex ◽  
Water Soluble ◽  
Large Hole ◽  
Membrane Pores ◽  
Model Lipid Membranes ◽  
Membrane Bound ◽  
Pore Forming Proteins

Pneumolysin is a member of the cholesterol-dependent cytolysin (CDC) family of pore-forming proteins that are produced as water-soluble monomers or dimers, bind to target membranes and oligomerize into large ring-shaped assemblies comprising approximately 40 subunits and approximately 30 nm across. This pre-pore assembly then refolds to punch a large hole in the lipid bilayer. However, in addition to forming large pores, pneumolysin and other CDCs form smaller lesions characterized by low electrical conductance. Owing to the observation of arc-like (rather than full-ring) oligomers by electron microscopy, it has been hypothesized that smaller oligomers explain smaller functional pores. To investigate whether this is the case, we performed cryo-electron tomography of pneumolysin oligomers on model lipid membranes. We then used sub-tomogram classification and averaging to determine representative membrane-bound low-resolution structures and identified pre-pores versus pores by the presence of membrane within the oligomeric curve. We found pre-pore and pore forms of both complete (ring) and incomplete (arc) oligomers and conclude that arc-shaped oligomeric assemblies of pneumolysin can form pores. As the CDCs are evolutionarily related to the membrane attack complex/perforin family of proteins, which also form variably sized pores, our findings are of relevance to that class of proteins as well.

Heterogeneity of High-molecular-weight Human Salivary Mucins

2000 ◽  
Vol 14 (1) ◽  
pp. 69-75 ◽  
Author(s):  
G.D. Offner ◽  
R.F. Troxler
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Biochemical Properties ◽  
Salivary Proteins ◽  
Molecular Weights ◽  
Structural Framework ◽  
Buccal Epithelial Cells ◽  
Membrane Bound ◽  
Membrane Spanning ◽  
Micro Organisms

The existence of high-molecular-weight glycoproteins in saliva and salivary secretions has been recognized for nearly 30 years. These proteins, called mucins, are essential for oral health and perform many diverse functions in the oral cavity. Mucins have been intensively studied, and much has been learned about their biochemical properties and their interactions with oral micro-organisms and other salivary proteins. In the past several years, the major high-molecular-weight mucin in salivary secretions has been identified as MUC5B, one of a family of 11 human mucin gene products expressed in tissue-specific patterns in the gastrointestinal, respiratory, and reproductive tracts. MUC5B is one of four gel-forming mucins which exist as multimeric proteins with molecular weights greater than 20-40 million daltons. The heavily glycosylated mucin multimers form viscous layers which protect underlying epithelial surfaces from microbial, mechanical, and chemical assault. Another class of mucin molecules, the membrane-bound mucins, is structurally and functionally distinct from the gel-forming mucins. These proteins do not form multimers and can exist as both secreted and membrane-bound forms, with the latter anchored to epithelial cell membranes through a short membrane-spanning domain. In the present work, we show that two of the membrane-bound mucins, MUC1 and MUC4, are expressed in all major human salivary glands as well as in buccal epithelial cells. While the functions of these mucins in the oral environment are not understood, it is possible that they form a structural framework on the cell surface which not only is cytoprotective, but also may serve as a scaffold upon which MUC5B, and possibly other salivary proteins, assemble.

scholarly journals Flip-flop asymmetry of cholesterol in model membranes induced by thermal gradients

Soft Matter ◽  
2020 ◽  
Vol 16 (25) ◽  
pp. 5925-5932
Author(s):  
James W. Carter ◽  
Miguel A. Gonzalez ◽  
Nicholas J. Brooks ◽  
John M. Seddon ◽  
Fernando Bresme
Keyword(s):  
Lipid Membranes ◽  
Model Membranes ◽  
Thermal Gradients ◽  
Flip Flop

Thermal gradients induce flip-flop asymmetry of cholesterol in lipid membranes.

scholarly journals Membrane proteins: always an insoluble problem?

2016 ◽  
Vol 44 (3) ◽  
pp. 790-795 ◽  
Author(s):  
Andrea E. Rawlings
Keyword(s):  
Membrane Proteins ◽  
Lipid Membranes ◽  
Drug Targets ◽  
Functional Characterization ◽  
Water Soluble ◽  
High Yield ◽  
Cellular Processes ◽  
Native Sequence ◽  
Protein Research ◽  
New Methodologies

Membrane proteins play crucial roles in cellular processes and are often important pharmacological drug targets. The hydrophobic properties of these proteins make full structural and functional characterization challenging because of the need to use detergents or other solubilizing agents when extracting them from their native lipid membranes. To aid membrane protein research, new methodologies are required to allow these proteins to be expressed and purified cheaply, easily, in high yield and to provide water soluble proteins for subsequent study. This mini review focuses on the relatively new area of water soluble membrane proteins and in particular two innovative approaches: the redesign of membrane proteins to yield water soluble variants and how adding solubilizing fusion proteins can help to overcome these challenges. This review also looks at naturally occurring membrane proteins, which are able to exist as stable, functional, water soluble assemblies with no alteration to their native sequence.

scholarly journals Catalase and Ascorbate Peroxidase in Euglenozoan Protists

Pathogens ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 317 ◽  
Author(s):  
Ingrid Škodová-Sveráková ◽  
Kristína Záhonová ◽  
Barbora Bučková ◽  
Zoltán Füssy ◽  
Vyacheslav Yurchenko ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Ascorbate Peroxidase ◽  
Biochemical Properties ◽  
Heterotrophic Flagellates ◽  
Oxygen Species ◽  
Detoxifying Enzymes ◽  
Free Living ◽  
Present Evidence ◽  
Obligate Parasites ◽  
Membrane Bound

In this work, we studied the biochemical properties and evolutionary histories of catalase (CAT) and ascorbate peroxidase (APX), two central enzymes of reactive oxygen species detoxification, across the highly diverse clade Eugenozoa. This clade encompasses free-living phototrophic and heterotrophic flagellates, as well as obligate parasites of insects, vertebrates, and plants. We present evidence of several independent acquisitions of CAT by horizontal gene transfers and evolutionary novelties associated with the APX presence. We posit that Euglenozoa recruit these detoxifying enzymes for specific molecular tasks, such as photosynthesis in euglenids and membrane-bound peroxidase activity in kinetoplastids and some diplonemids.

Water-Soluble Conjugated Polymers for Fluorescent-Enzyme Assays

2010 ◽  
Vol 31 (16) ◽  
pp. 1405-1421 ◽  
Author(s):  
Fude Feng ◽  
Libing Liu ◽  
Qiong Yang ◽  
Shu Wang
Keyword(s):  
Conjugated Polymers ◽  
Water Soluble ◽  
Enzyme Assays

scholarly journals Ladderane phospholipids form a densely packed membrane with normal hydrazine and anomalously low proton/hydroxide permeability

2018 ◽  
Vol 115 (37) ◽  
pp. 9098-9103 ◽  
Author(s):  
Frank R. Moss ◽  
Steven R. Shuken ◽  
Jaron A. M. Mercer ◽  
Carolyn M. Cohen ◽  
Thomas M. Weiss ◽  
...  
Keyword(s):  
Lipid Bilayers ◽  
Lipid Membranes ◽  
Water Soluble ◽  
Anammox Bacteria ◽  
Straight Chain ◽  
Slow Growth Rate ◽  
Hydrazine Sensor ◽  
Diffusion Assay ◽  
Ladderane Lipids ◽  
Transmembrane Diffusion

Ladderane lipids are unique to anaerobic ammonium-oxidizing (anammox) bacteria and are enriched in the membrane of the anammoxosome, an organelle thought to compartmentalize the anammox process, which involves the toxic intermediate hydrazine (N2H4). Due to the slow growth rate of anammox bacteria and difficulty of isolating pure ladderane lipids, experimental evidence of the biological function of ladderanes is lacking. We have synthesized two natural and one unnatural ladderane phosphatidylcholine lipids and compared their thermotropic properties in self-assembled bilayers to distinguish between [3]- and [5]-ladderane function. We developed a hydrazine transmembrane diffusion assay using a water-soluble derivative of a hydrazine sensor and determined that ladderane membranes are as permeable to hydrazine as straight-chain lipid bilayers. However, pH equilibration across ladderane membranes occurs 5–10 times more slowly than across straight-chain lipid membranes. Langmuir monolayer analysis and the rates of fluorescence recovery after photobleaching suggest that dense ladderane packing may preclude formation of proton/hydroxide-conducting water wires. These data support the hypothesis that ladderanes prevent the breakdown of the proton motive force rather than blocking hydrazine transmembrane diffusion in anammox bacteria.

Delivery of Active FGF-2 From Mechanically-Stable Biological Nanofibers Accelerates Cell Ingress Into Multifiber Composites

2011 ◽  
Author(s):  
Jonathan A. Kluge ◽  
Rudra A. Pampati ◽  
Mara L. Schenker ◽  
Daniel J. Zhou ◽  
John E. Esterhai ◽  
...  
Keyword(s):  
Release Kinetics ◽  
Composite Fiber ◽  
Biochemical Properties ◽  
Fiber Spinning ◽  
Water Soluble ◽  
Silk Fibers ◽  
Synthetic Fibers ◽  
Nanofibrous Scaffolds ◽  
Biologic Factors

Fibrocartilaginous tissues such as the meniscus and annulus fibrosus serve critical load-bearing roles, relying on arrays of highly organized collagen fibers to resist tensile loads [1]. As these specialized structures are often injured, there exists great demand for engineered tissues for repair or replacement. Cell-laden aligned nanofibrous scaffolds formed from poly(ε-caprolactone) (PCL) have shown promise in achieving tissuelike mechanical and biochemical properties and can direct cellular and matrix organization in vitro [2]. A current limitation of nanofibrous scaffolds, however, is a slow rate of cellular infiltration, particularly in thick scaffolds. To address this, dynamic composite nanofibrous scaffolds have been fabricated via multi-fiber spinning [3], which can offer tunable modes of degradation depending on the polymer sources. For example, water-soluble polyethylene oxide (PEO) fibers can be co-spun with PCL to improve porosity and hasten cell ingress [4]. Incorporation of additional tunable and bioactive polymer sources may add greater versatility to these composite systems. For example, aqueous-based silk fibroin can be used as a slow-degrading, mechanically strong composite fiber component [5] into which active biologic factors (drugs, growth factors) can be incorporated [6]. Variably-degradable silk fibers can be formed by modulating post-spinning treatments, and protein release kinetics can likewise be manipulated by the physical crosslinking method [7]. We hypothesized that incorporation of robust and tunable silk protein-based fibers into a composite of slow-degrading synthetic fibers would provide mechanical function while delivering active biologic factors to expedite cell proliferation and encourage more rapid construct colonization. To test this hypothesis, we characterized the release kinetics of recombinant FGF-2 from silk fibers and its bioactivity in vitro and in a rat subcutaneous implant model.

Oxyfluorfen and Lipid Peroxidation: Protein Damage as a Phytotoxic Consequence

Weed Science ◽  
1985 ◽  
Vol 33 (6) ◽  
pp. 766-770 ◽  
Author(s):  
Karl J. Kunert ◽  
Carmen Homrighausen ◽  
Herbert Böhme ◽  
Peter Böger
Keyword(s):  
Lipid Peroxidation ◽  
Cytochrome C ◽  
Diphenyl Ether ◽  
Photosynthetic Electron Transport ◽  
Water Soluble ◽  
Cytochrome F ◽  
Protein Damage ◽  
Protein Loss ◽  
Membrane Bound

Protein damage, as a primary phytotoxic consequence of in vivo lipid peroxidation, induced by the diphenyl ether herbicide oxyfluorfen [2-chloro-1-(3-ethoxy-4-nitrophenoxy)-4-(trifluoromethyl)benzene] at a concentration of 10 μM, was measured with the green algaScenedesmus acutus. In the light, water-soluble proteins are destroyed by a herbicide-induced peroxidation process that can be detected by production of fluorescent products and loss of specific amino acid residues of proteins. The water-soluble cytochrome c-553 and the membrane-bound cytochrome f-553, components of the photosynthetic electron transport, were specifically used as sensitive markers for protein damage, measured as decrease of redox reactions of the cytochromes. Under peroxidizing conditions, destruction of the algal cytochrome c is significantly higher than destruction of membrane-bound components, such as cytochrome f and chlorophyll. Protection against protein loss is achieved by the nonbiological antioxidant ethoxyquin (1,2-dihydro-6-ethoxy-2,2,4-trimethylquinoline) or the photosynthesis inhibitor diuron [N′-(3,4-dichlorophenyl)-N,N-dimethylurea].

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