Tilted peptides: a structural motif involved in protein membrane insertion?

Tilted peptides are short hydrophobic protein fragments characterized by an asymmetric distribution of their hydrophobic residues when helical. They are able to interact with a hydrophobic/hydrophilic interface (such as a lipid membrane) and to destabilize the organized system into which they insert. They were detected in viral fusion proteins and in proteins involved in different biological processes involving membrane insertion or translocation of the protein in which they are found. In this paper, we have analysed different protein domains related to membrane insertion with regard to their tilted properties. They are the N-terminal signal peptide of the filamentous haemagglutinin (FHA), a Bordetella pertussis protein secreted in high amount and the hydrophobic domain from proteins forming pores (i.e. ColIa, Bax and Bcl-2). From the predictions and the experimental approaches, we suggest that tilted peptides found in those proteins could have a more general role in the mechanism of insertion/translocation of proteins into/across membranes. For the signal sequences, they could help the protein machinery involved in protein secretion to be more active. In the case of toroidal pore formation, they could disturb the lipids, facilitating the insertion of the other more hydrophilic helices.     

Chromatin globules: a common motif of higher order chromosome structure?

Recent technological advances in the field of chromosome conformation capture are facilitating tremendous progress in the ability to map the three-dimensional (3D) organization of chromosomes at a resolution of several Kb and at the scale of complete genomes. Here we review progress in analyzing chromosome organization in human cells by building 3D models of chromatin based on comprehensive chromatin interaction datasets. We describe recent experiments that suggest that long-range interactions between active functional elements are sufficient to drive folding of local chromatin domains into compact globular states. We propose that chromatin globules are commonly formed along chromosomes, in a cell type specific pattern, as a result of frequent long-range interactions among active genes and nearby regulatory elements. Further, we speculate that increasingly longer range interactions can drive aggregation of groups of globular domains. This process would yield a compartmentalized chromosome conformation, consistent with recent observations obtained with genome-wide chromatin interaction mapping.     

Inheritance of the replication complex: a unique or common phenomenon in the control of DNA replication?

Early models of the regulation of initiation of DNA replication by protein complexes predicted that binding of a replication initiator protein to a replicator region is required for initiation of each DNA replication round, since after the initiation event the replication initiator should dissociate from DNA. It was, therefore, assumed that binding of the replication initiator is a signal for triggering DNA replication. However, more recent investigations have revealed that in many replicons this is not the case. Studies on the regulation of the replication of plasmids derived from bacteriophage lambda demonstrated that, once assembled, the replication complex can be inherited by one of the two daughter plasmid copies after each replication round and may function in subsequent replication rounds. Since this DNA-bound protein complex bears information about specific initiation of DNA replication, this phenomenon has been called "protein inheritance." A similar phenomenon has recently been reported for oriJ-based plasmids. Moreover, the current model of the initiation of DNA replication in the yeast Saccharomyces cerevisiae proposes that the origin recognition complex (ORC) remains bound to one copy of the ori sequence (the ARS region) after initiation of DNA replication. Thus, it seems plausible that protein inheritance is not unique for lambda plasmids, but may be a common phenomenon in the control of DNA replication, at least in microbes.     

Phylloxera and the grapevine: a sense of common purpose?

The purpose of life is its continuation: survival is the reason things live. Here we explore this 'basic' of biology, by reference to the extraordinary life-cycle of the aphid-like pest phylloxera, and the complexity of its relationship with its host the grapevine. The effort and ingenuity that phylloxera employs to continue itself leads to a doubt that survival alone is sufficient reason. It has frequently been suggested that the reduction of life to a catalogue of facts (by science) creates this doubt, because it robs existence of its essence (which is something other than its mechanics). The part that science is said to steal is what Robert Pirsig calls Quality-the harmonious balance of things. Pirsig seems to imply that this is something inherent in things-and independent from us. A more mundane explanation is that the difference between facts and the complete reality is us-the tendency of mind to connect freely between different kinds of information. This possibility is briefly illustrated here by a myth based on the facts of phylloxera.     

Reactivity of glycoconjugate in membrane system II: Can a neutral glycolipid function as lectin receptor in the presence of gangliosides in plasma membrane?

To examine how surface Potential controls the reactivity of glycoconjugates at cell surface, the interaction of galactose-sPecific lectinse.g. peanut agglutinin,Ricinus cummunis agglutinin with liPosomes bearing asialo GM₁ were studied in the Presence of varying amount of ganglioside mixture, GM_n. The Presence of 5% GM_n causes comPlete slowing down of PreciPitin reaction and thereby make carbohydrate moiety of asialo GM₁ comPletely inaccessiblei.e. ‘cryPtic’. In contrast the Presence of 1–2% GM_n enhances the aPParent rate and amPlitude of the PreciPitin reaction as surface Potential becomes more negative. The relevance of the findings has been discussed in relation to the exPression and involvement of the cell-surface sialic acid residues during develoPment and differentiation.     

Is the WKS motif the tissue-factor binding site for coagulation factor VII?

Human tissue factor (TF), the membrane-bound glycoprotein receptor for the blood-clotting factor VII/VIIa, contains in its extracellular domain three repeats of the rare motif, tryptophan-lysine-serine (WKS). Murine tissue factor, which binds human factor VII/VIIa poorly, contains only one WKS motif suggesting that the WKS motif may be involved in the binding of human factor VII/VIIa to human TF. Sequence analysis has revealed a WKS motif in 23 human proteins, seven of which are involved in the coagulation process. Another five WKS-containing proteins share some functional properties with the coagulation proteins. Analysis of the properties of these proteins provides some insight into the possible functional role of the WKS motif.     

Oxidative degradation of cationic metalloporphyrins in the presence of nucleic acids: a way to binding constants?

Mn(III) and Fe(III) complexes of meso-tetrakis(N-methylpyridinium-4-yl)porphyrin (M-TMePyP) and related hybrid molecules ("metalloporphyrin-ellipticine") were activated by potassium monopersulfate in the presence of variable calf thymus (CT) DNA and NaCl concentrations. Monitored by visible spectroscopy (Soret band), fast degradation of the free metalloprophyrin was observed while the DNA-bound form appeared protected. This direct quantitation of free versus bound metalloporphyrin ratios allowed determination of binding constants: Mn- and Fe-TMePyP respectively bind to CT DNA (5 mM phosphate buffer, 0.1 M NaCl, pH 7) with K = 3 X 10(4) and 1.2 X 10(4) M-1. Mn-TMePyP showed a greater affinity for poly[d(A-T)] (K = 1.2 X 10(5) M-1) than for poly[d(G-C)] (K = 0.2 X 10(4) M-1). This method allowed us access to the intrinsic DNA affinity of the metalloporphyrin moiety of the hybrid molecules "metalloporphyrin-ellipticine".     

Locally resolved membrane binding affinity of the N-terminus of α-synuclein

α-Synuclein is abundantly present in Lewy bodies, characteristic of Parkinson's disease. Its exact physiological role has yet to be determined, but mitochondrial membrane binding is suspected to be a key aspect of its function. Electron paramagnetic resonance spectroscopy in combination with site-directed spin labeling allowed for a locally resolved analysis of the protein-membrane binding affinity for artificial phospholipid membranes, supported by a study of binding to isolated mitochondria. The data reveal that the binding affinity of the N-terminus is nonuniform.     

Clathrin-binding proteins: got a motif? Join the network!

Clathrin plays a key function in membrane and protein trafficking through the endocytic and late secretory pathways. Its role as a molecular scaffold that drives formation of transport vesicles requires binding to a number of proteins with distinct functional and structural properties. Recent studies have revealed that most of these proteins interact with clathrin through surprisingly simple, linear arrangements of acidic and hydrophobic amino acid residues. This article discusses the different types of clathrin-binding proteins and motifs as well as the physiological significance of these proteins in clathrin-dependent events.     

Individual differences in the chemical senses: is there a common sensitivity?

Taste, smell, and chemical irritation (so-called trigeminal sensation) combine in our daily experience to produce the supramodal sensation of flavor, are processed by partly overlapping neural mechanisms, and show functional interconnectivity in experiments. Given their collaboration in flavor formation and the well-established connections between these senses, it is plausible that polymodal detection mechanisms might contribute to individual differences in measured sensitivity. One would expect the existence of a general chemosensory sensitivity factor to result in associations among taste, smell, and trigeminal stimulation thresholds. Measures of 5 detection thresholds from all the chemical senses were assessed in the same group of young healthy subjects (n=57). An unbiased principal components analysis (PCA) yielded a 2-component solution. Component 1, on which taste thresholds loaded strongly, accounted for 29.4% of the total variance. Component 2, on which the odor and trigeminal lateralization thresholds loaded strongly, accounted for 26.9% of the total variance. A subsequent PCA restricted to a 3-component solution cleanly separated the 3 sensory modalities and accounted for 75% of the total variance. Thus, though there may be a common underlying factor that determines some individual differences in odor and trigeminal lateralization thresholds, a general chemical sensitivity that spans chemosensory modalities seems unlikely.     

Staufen: a common component of mRNA transport in oocytes and neurons?

Mammalian homologues of Staufen, a protein involved in localizing mRNAs during oogenesis and early central nervous system development in Drosophila, have been identified recently. The mammalian staufen gene encodes a protein containing several conserved double-stranded mRNA-binding domains and is expressed in hippocampal neurons. The mammalian Staufen protein forms granules that are transported to the distal dendrite during neuronal maturation. The Staufen granules colocalize with ribonuclear particles that transport mRNA to the dendrites. These findings might provide clues to a mechanism of mRNA transport conserved in mammalian neurons and Drosophila oogenesis.     

Cys-labeling kinetics of membrane protein GlpG: a role for specific SDS binding and micelle changes?

Empirically, α-helical membrane protein folding stability in surfactant micelles can be tuned by varying the mole fraction MF^SDS of anionic (sodium dodecyl sulfate (SDS)) relative to nonionic (e.g., dodecyl maltoside (DDM)) surfactant, but we lack a satisfying physical explanation of this phenomenon. Cysteine labeling (CL) has thus far only been used to study the topology of membrane proteins, not their stability or folding behavior. Here, we use CL to investigate membrane protein folding in mixed DDM-SDS micelles. Labeling kinetics of the intramembrane protease GlpG are consistent with simple two-state unfolding-and-exchange rates for seven single-Cys GlpG variants over most of the explored MF^SDS range, along with exchange from the native state at low MF^SDS (which inconveniently precludes measurement of unfolding kinetics under native conditions). However, for two mutants, labeling rates decline with MF^SDS at 0–0.2 MF^SDS (i.e., native conditions). Thus, an increase in MF^SDS seems to be a protective factor for these two positions, but not for the five others. We propose different scenarios to explain this and find the most plausible ones to involve preferential binding of SDS monomers to the site of CL (based on computational simulations) along with changes in size and shape of the mixed micelle with changing MF^SDS (based on SAXS studies). These nonlinear impacts on protein stability highlights a multifaceted role for SDS in membrane protein denaturation, involving both direct interactions of monomeric SDS and changes in micelle size and shape along with the general effects on protein stability of changes in micelle composition.     

Is lipid bilayer binding a common property of inhibitor cysteine knot ion-channel blockers?

Recent studies of several ICK ion-channel blockers suggest that lipid bilayer interactions play a prominent role in their actions. Structural similarities led to the hypothesis that bilayer interactions are important for the entire ICK family. We have tested this hypothesis by performing direct measurements of the free energy of bilayer partitioning (DeltaG) of several peptide blockers using our novel quenching-enhanced fluorescence titration protocol. We show that various ICK peptides demonstrate markedly different modes of interaction with large unilamellar lipid vesicles. The mechanosensitive channel blocker, GsMTx4, and its active diastereomeric analog, D-GsMTx4, bind strongly to both anionic and zwitterionic membranes. One potassium channel gating modifier, rHpTx2gs, interacts negligibly with both types of vesicles at physiological pH, whereas another, SGTx1, interacts only with anionic lipids. The slope of DeltaG dependence on surface potential is very shallow for both GsMTx4 and D-GsMTx4, indicating complex interplay of their hydrophobic and electrostatic interactions with lipid. In contrast, a cell-volume regulator, GsMTx1, and SGTx1 exhibit a very steep DeltaG dependence on surface potential, resulting in a strong binding only for membranes rich in anionic lipids. The high variability of 5 kcal/mole in observed DeltaG shows that bilayer partitioning is not a universal property of the ICK peptides interacting with ion channels.     

Is the isolated ligand binding domain a good model of the domain in the native receptor?

Numerous studies have used the atomic level structure of the isolated ligand binding domain of the glutamate receptor to elucidate the agonist-induced activation and desensitization processes in this group of proteins. However, no study has demonstrated the structural equivalence of the isolated ligand binding fragments and the protein in the native receptor. In this report, using visible absorption spectroscopy we show that the electronic environment of the antagonist 6-cyano-7-nitro-2,3-dihydroxyquinoxaline is identical for the isolated protein and the native glutamate receptors expressed in cells. Our results hence establish that the local structure of the ligand binding site is the same in the two proteins and validate the detailed structure-function relationships that have been developed based on a comparison of the structure of the isolated ligand binding domain and electrophysiological consequences in the native receptor.     

Terminal disorder: a common structural feature of the axial proteins of bacterial flagellum?

We report, based on proteolytic experiments and high resolution 1H nuclear magnetic resonance studies that the terminal regions of the monomeric hook protein are highly mobile and exposed to the solvent. The disordered parts of the hook protein span approximately the first 70 and the last 30 amino acid residues. Although the amino acid sequences of flagellin and hook protein do not resemble each other at all, both proteins have now been shown to contain large disordered terminal regions. Sequential similarities of flagellin and hook protein, especially near the NH2 and COOH termini, to other axial components of bacterial flagellum suggest that terminal disorder may be a common structural feature of the axial proteins of the bacterial flagellum.     

New insights into the role of mast cells in autoimmunity: Evidence for a common mechanism of action?

Mast cells are classically considered innate immune cells that act as first responders in many microbial infections and have long been appreciated as potent contributors to allergic reactions. However, recent advances in the realm of autoimmunity have made it clear that these cells are also involved in the pathogenic responses that exacerbate disease. In the murine models of multiple sclerosis, rheumatoid arthritis and bullous pemphigoid, both the pathogenic role of mast cells and some of their mechanisms of action are shared. Similar to their role in infection and a subset of allergic responses, mast cells are required for the efficient recruitment of neutrophils to sites of inflammation. Although this mast cell-dependent neutrophil response is protective in infection settings, it is postulated that neutrophils promote local vascular permeability and facilitate the entry of inflammatory cells that enhance tissue destruction at target sites. However, there is still much to learn. There is little information regarding mechanisms of mast cell activation in disease. Nor is it known how many mast cell-derived mediators are relevant and whether interactions with other cells are implicated in these diseases including T cells, B cells and astrocytes. Here we review the current state of knowledge about mast cells in autoimmune disease. We also discuss findings regarding newly discovered mast cell actions and factors that modulate mast cell function. We speculate that much of this new information will ultimately contribute to a greater understanding of the full range of mast cell actions in autoimmunity. This article is part of a Special Issue entitled: Mast cells in inflammation.