Interactions of the Lysosomotropic Detergent O-Methyl-Serine Dodecylamide Hydrochloride (MSDH) with Lipid Bilayer Membranes-Implications for Cell Toxicity.

O-methyl-serine dodecylamine hydrochloride (MSDH) is a detergent that accumulates selectively in lysosomes, a so-called lysosomotropic detergent, with unexpected chemical properties. At physiological pH, it spontaneously forms vesicles, which disassemble into small aggregates (probably micelles) below pH 6.4.

Lipid vesicles affect the aggregation of 4-hydroxy-2-nonenal-modified α-synuclein oligomers.

Parkinson's disease (PD) and other synucleinopathies are characterized by accumulation of misfolded aggregates of α-synuclein (α-syn). The normal function of α-syn is still under investigation, but it has been generally linked to synaptic plasticity, neurotransmitter release and the maintenance of the synaptic pool. α-Syn localizes at synaptic terminals where it can bind to synaptic vesicles as well as to other cellular membranes.

Impact of high cholesterol in a Parkinson's disease model: Prevention of lysosomal leakage versus stimulation of α-synuclein aggregation.

Parkinson's disease is characterized by accumulation of intraneuronal cytoplasmic inclusions, Lewy bodies, which mainly consist of aggregated α-synuclein. Controversies exist as to whether high blood cholesterol is a risk factor for the development of the disease and whether statin treatment could have a protective effect. Using a model system of BE(2)-M17 neuroblastoma cells treated with the neurotoxin 1-methyl-4-phenylpyridinium (MPP), we found that MPP-induced cell death was accompanied by cholesterol accumulation in a lysosomal-like pattern in pre-apoptotic cells.

VirB6 and VirB10 from the Brucella type IV secretion system interact via the N-terminal periplasmic domain of VirB6.

Type IV secretion systems are multi-protein complexes that transfer macromolecules across the cell envelope of bacteria. Identifying the sites of interaction between the twelve proteins (VirB1-VirB11 and VirD4) that form these complexes is key to understanding their assembly and function. We have here used phage display, bacterial two-hybrid and fluorescence-based interaction assays to identify an N-terminal domain of the inner membrane protein VirB6 as a site of interaction with the envelope-spanning VirB10 protein.

Type IV secretion system core component VirB8 from Brucella binds to the globular domain of VirB5 and to a periplasmic domain of VirB6.

Type IV secretion systems are macromolecular assemblies in the cell envelopes of bacteria that function in macromolecular translocation. Structural biology approaches have provided insights into the interaction of core complex components, but information about proteins that undergo transient interactions with membrane components has not been forthcoming. We have pursued an unbiased approach using peptide arrays and phage display to identify interaction partners and interaction domains of type IV secretion system assembly factor VirB8.

Plasma membrane calcium pump (PMCA) differential exposure of hydrophobic domains after calmodulin and phosphatidic acid activation.

The exposure of the plasma membrane calcium pump (PMCA) to the surrounding phospholipids was assessed by measuring the incorporation of the photoactivatable phosphatidylcholine analog [(125)I]TID-PC/16 to the protein. In the presence of Ca(2+) both calmodulin (CaM) and phosphatidic acid (PA) greatly decreased the incorporation of [(125)I]TID-PC/16 to PMCA. Proteolysis of PMCA with V8 protease results in three main fragments: N, which includes transmembrane segments M1 and M2; M, which includes M3 and M4; and C, which includes M5 to M10.

Quantitative analysis of VirB8-VirB9-VirB10 interactions provides a dynamic model of type IV secretion system core complex assembly.

Type IV secretion systems are multiprotein complexes that translocate macromolecules across the bacterial cell envelope. The type IV secretion system in Brucella species encodes 12 VirB proteins that permit this pathogen to translocate effectors into mammalian cells, where they contribute to its survival inside the host. The "core" complex proteins are conserved in all type IV secretion systems, and they are believed to form the channel for substrate translocation.

The structure of calreticulin C-terminal domain is modulated by physiological variations of calcium concentration.

Calreticulin is an abundant endoplasmic reticulum resident protein that fulfills at least two basic functions. Firstly, due to its ability to bind monoglucosylated high mannose oligosaccharides, calreticulin is a central component of the folding quality control system of glycoproteins. On the other hand, thanks to its capacity to bind high amounts of calcium, calreticulin is one of the main calcium buffers in the endoplasmic reticulum.

A new conformation in sarcoplasmic reticulum calcium pump and plasma membrane Ca2+ pumps revealed by a photoactivatable phospholipidic probe.

The purpose of this work was to obtain structural information about conformational changes in the membrane region of the sarcoplasmic reticulum (SERCA) and plasma membrane (PMCA) Ca(2+) pumps. We have assessed changes in the overall exposure of these proteins to surrounding lipids by quantifying the extent of protein labeling by a photoactivatable phosphatidylcholine analog 1-palmitoyl-2-[9-[2'-[(125)I]iodo-4'-(trifluoromethyldiazirinyl)-benzyloxycarbonyl]-nonaoyl]-sn-glycero-3-phosphocholine ([(125)I]TID-PC/16) under different conditions. We determined the following.

Phospholipid distribution around the plasma membrane calcium pump: a hydrophobic photolabeling study.

The functions of membrane proteins are highly dependent on their phospholipid environment. In this article, we have used a hydrophobic photolabeling method to study the noncovalent interactions between plasma membrane calcium pump (PMCA) and surrounding phospholipids. With this approach, we determined (1) the number of lipid molecules in close contact with the transmembrane surface, i.

Stoichiometry of lipid-protein interaction assessed by hydrophobic photolabeling.

Here we undertook a comparative study of the composition of the lipid annulus of three ATPases pertaining to the P-type family: plasma membrane calcium pump (PMCA), sarcoplasmic reticulum calcium pump (SERCA) and Na,K-ATPase. The photoactivatable phosphatidylcholine analogue [(125)I]TID-PC/16 was incorporated into mixtures of dimyristoyl phosphatidylcholine (DMPC) and each enzyme with the aid of the nonionic detergent C(12)E(10). After photolysis, the extent of the labeling reaction was assessed to determine the lipid:protein stoichiometry: 17 for PMCA, 18 for SERCA, 24 for the Na,K-ATPase (alpha-subunit) and 5.