scholarly journals Membrane Binding by MinD Involves Insertion of Hydrophobic Residues within the C-Terminal Amphipathic Helix into the Bilayer

2003 ◽  
Vol 185 (15) ◽  
pp. 4326-4335 ◽  
Author(s):  
Huaijin Zhou ◽  
Joe Lutkenhaus
Keyword(s):  
Emission Spectroscopy ◽  
Membrane Binding ◽  
Fluorescence Emission ◽  
Phospholipid Vesicles ◽  
Amphipathic Helix ◽  
Spectroscopy Analysis ◽  
Membrane Bilayer ◽  
Hydrophobic Residues ◽  
Tryptophan Residues ◽  
The Mind

ABSTRACT MinD binds to phospholipid vesicles in the presence of ATP and is released by MinE, which stimulates the MinD ATPase. Membrane binding requires a short conserved C-terminal region, which has the potential to form an amphipathic helix. This finding has led to a model in which the binding of ATP regulates the formation or accessibility of this helix, which then embeds in the membrane bilayer. To test this model, we replaced each of the four hydrophobic residues within this potential helix with tryptophan or a charged residue. Introduction of a negatively charged amino acid decreased membrane binding of MinD and its ability to activate MinC. In contrast, mutants with tryptophan substitutions retained the ability to bind to the membrane and activate MinC. Fluorescence emission spectroscopy analysis of the tryptophan mutants F263W, L264W, and L267W confirmed that these tryptophan residues did insert into the hydrophobic interior of the bilayer. We conclude that membrane binding by MinD involves penetration of the hydrophobic residues within the C-terminal amphipathic helix into the hydrophobic interior of the bilayer.

scholarly journals A Fast, Straightforward and Inexpensive Method for the Authentication of Baijiu Spirit Samples by Fluorescence Spectroscopy

Beverages ◽  
2021 ◽  
Vol 7 (3) ◽  
pp. 65
Author(s):  
Rachel L. Burns ◽  
Raegan Alexander ◽  
Liliya Snaychuk ◽  
John C. Edwards ◽  
Neil Fitzgerald ◽  
...  
Keyword(s):  
Analytical Chemistry ◽  
North America ◽  
Fluorescence Spectroscopy ◽  
Emission Spectroscopy ◽  
Low Cost ◽  
Fluorescence Emission ◽  
Wide Distribution ◽  
Inexpensive Method ◽  
Scientific Methods ◽  
Price Differences

The Chinese spirit baijiu is currently the world’s bestselling spirit, with more than ten billion liters sold in 2018. This is a figure that puts its sales higher than whiskey, vodka, gin, and tequila combined. The multitude of baijiu varieties available in the market differ in several ways ranging from aging to the traditional artisanship involved in producing the final spirit to several other features, including the rarity of the bottle. A result of these differences is a wide distribution of prices for the various baijiu products. Consequently, a single bottle of baijiu can cost anywhere from a few dollars, up to thousands of US dollars. The price differences among the various baijiu spirits necessitate the existence of reliable scientific methods that can efficiently differentiate and authenticate the qualities of baijiu spirits. In addition, the existence of such methods facilitates the prevention of counterfeit sales of the final product. Considering this, we introduce an analytical chemistry method that distinguishes amongst different baijiu spirits based on fluorescence spectroscopy. Its attributes include the low cost and convenience that allows analysis either before or while the spirit is in the market. Our work herein focuses on the analysis of thirty different varieties of baijiu spirits from six different distilleries from East Asia and North America by fluorescence emission spectroscopy, which is associated to the price of the product. For the analysis, we employed a HORIBA FLUOROLOG 3 (HORIBA—Jobin Yvon) spectrometer. Major advantages of this method include the low cost, as no consumables except a quartz reusable cuvette are required, the minimal waste, and finally the quick processing of data.

Role of Cationic Groove and Hydrophobic Residues in Phosphatidylinositol-Dependent Membrane-Binding Properties of Tks5-Phox Homology Domain

2018 ◽  
Vol 3 (4) ◽  
pp. 1205-1214 ◽  
Author(s):  
Sukhamoy Gorai ◽  
Debasish Paul ◽  
Rituparna Borah ◽  
Nandan Haloi ◽  
Manas Kumar Santra ◽  
...  
Keyword(s):  
Membrane Binding ◽  
Binding Properties ◽  
Hydrophobic Residues ◽  
Phox Homology

scholarly journals Benzoate Decreases the Binding of cis,cis-Muconate to the BenM Regulator despite the Synergistic Effect of Both Compounds on Transcriptional Activation

2004 ◽  
Vol 186 (4) ◽  
pp. 1200-1204 ◽  
Author(s):  
Todd J. Clark ◽  
Robert S. Phillips ◽  
Becky M. Bundy ◽  
Cory Momany ◽  
Ellen L. Neidle
Keyword(s):  
Dna Binding ◽  
Synergistic Effect ◽  
Dissociation Constant ◽  
Transcriptional Activation ◽  
Emission Spectroscopy ◽  
Fluorescence Emission ◽  
Transcriptional Regulator ◽  
Dna Binding Domain ◽  
Apparent Dissociation Constant ◽  
High Level

ABSTRACT Fluorescence emission spectroscopy was used to investigate interactions between two effectors and BenM, a transcriptional regulator of benzoate catabolism. BenM had a higher affinity for cis,cis-muconate than for benzoate as the sole effector. However, the presence of benzoate increased the apparent dissociation constant (reduced the affinity) of the protein for cis,cis-muconate. Similar results were obtained with truncated BenM lacking the DNA-binding domain. High-level transcriptional activation may require that some monomers within a BenM tetramer bind benzoate and others bind cis,cis-muconate.

scholarly journals Toward Elucidating the Membrane Topology of Helix Two of the Colicin E1 Channel Domain

2006 ◽  
Vol 281 (43) ◽  
pp. 32375-32384 ◽  
Author(s):  
Dawn White ◽  
Abdiwahab A. Musse ◽  
Jie Wang ◽  
Erwin London ◽  
A. Rod Merrill
Keyword(s):  
Fluorescence Emission ◽  
Fluorescence Labeling ◽  
Interfacial Region ◽  
Closed Channel ◽  
Membrane Bilayer ◽  
Colicin E1 ◽  
Aqueous Solvent ◽  
Least Squares Fit ◽  
Membrane Bound ◽  
Α Helix

The membrane-bound closed state of the colicin E1 channel domain was investigated by site-directed fluorescence labeling using a bimane fluorophore attached to each single cysteine residue within helix 2 of each mutant protein. The fluorescence properties of the bimane fluorophore were measured for the membrane-associated form of the closed channel and included fluorescence emission maximum, fluorescence anisotropy, apparent polarity, surface accessibility, and membrane bilayer penetration depth. The fluorescence data show that helix 2 is an amphipathic α-helix that is situated parallel to the membrane surface, but it is less deeply embedded within the bilayer interfacial region than is helix 1 in the closed channel. A least squares fit of the various data sets to a harmonic wave function indicated that the periodicity and angular frequency for helix 2 in the membrane-bound state are typical for an amphipathic α-helix (3.8 ± 0.1 residues per turn and 94 ± 4°, respectively) that is located at an interfacial region of a membrane bilayer. Dual quencher analysis also revealed that helix 2 is peripherally membrane associated, with one face of the helix dipping into the interfacial region of the lipid bilayer and the other face projecting outwardly into the aqueous solvent. Finally, our data show that helices 1 and 2 remain independent helices upon membrane association with a short connector link (Tyr363–Gly364) and that short amphipathic α-helices participate in the formation of a lipid-dependent, toroidal pore for this colicin.

scholarly journals The conformational changes of α2-macroglobulin induced by methylamine or trypsin. Characterization by extrinsic and intrinsic spectroscopic probes

1987 ◽  
Vol 243 (1) ◽  
pp. 47-54 ◽  
Author(s):  
L J Larsson ◽  
P Lindahl ◽  
C Hallén-Sandgren ◽  
I Björk
Keyword(s):  
Conformational Change ◽  
Conformational Changes ◽  
Fluorescence Emission ◽  
Tryptophan Fluorescence ◽  
Cross Linking ◽  
Bait Region ◽  
Close Proximity ◽  
Tryptophan Residues ◽  
Thioester Bond ◽  
Bond Region

The conformational changes around the thioester-bond region of human or bovine alpha 2M (alpha 2-macroglobulin) on reaction with methylamine or trypsin were studied with the probe AEDANS [N-(acetylaminoethyl)-8-naphthylamine-1-sulphonic acid], bound to the liberated thiol groups. The binding affected the fluorescence emission and lifetime of the probe in a manner indicating that the thioester-bond region is partially buried in all forms of the inhibitor. In human alpha 2M these effects were greater for the trypsin-treated than for the methylamine-treated inhibitor, which both have undergone similar, major, conformational changes. This difference may thus be due to a close proximity of the thioester region to the bound proteinase. Reaction of trypsin with thiol-labelled methylamine-treated bovine alpha 2M, which retains a near-native conformation and inhibitory activity, indicated that the major conformational change accompanying the binding of proteinases involves transfer of the thioester-bond region to a more polar environment without increasing the exposure of this region at the surface of the protein. Labelling of the transglutaminase cross-linking site of human alpha 2M with dansylcadaverine [N-(5-aminopentyl)-5-dimethylaminonaphthalene-1-sulphonamide] suggested that this site is in moderately hydrophobic surroundings. Reaction of the labelled inhibitor with methylamine or trypsin produced fluorescence changes consistent with further burial of the cross-linking site. These changes were more pronounced for trypsin-treated than for methylamine-treated alpha 2M, presumably an effect of the cleavage of the adjacent ‘bait’ region. Solvent perturbation of the u.v. absorption and iodide quenching of the tryptophan fluorescence of human alpha 2M showed that one or two tryptophan residues in each alpha 2M monomer are buried on reaction with methylamine or trypsin, with no discernible change in the exposure of tyrosine residues. Together, these results indicate an extensive conformational change of alpha 2M on reaction with amines or proteinases and are consistent with several aspects of a recently proposed model of alpha 2M structure [Feldman, Gonias & Pizzo (1985) Proc. Natl. Acad. Sci. U.S.A. 82, 5700-5704].

scholarly journals A Study of Local Anaesthetics. Part 202. Determination of the Critical Micellar Concentration of Carbisocainium Chloride in Water Using Spectral Methods and the Probe Pyrene

2013 ◽  
Vol 60 (1) ◽  
pp. 1-6 ◽  
Author(s):  
J. Gališinová ◽  
F. Andriamainty ◽  
I. Malík ◽  
J. Čižmárik ◽  
J. Karlovská ◽  
...  
Keyword(s):  
Spectral Methods ◽  
Emission Spectroscopy ◽  
Surfactant Concentration ◽  
Fluorescence Emission ◽  
Local Anaesthetics ◽  
Critical Micellar Concentration ◽  
Ratio Data ◽  
Micellization Process ◽  
Second Procedure

The micellization process of the local anaesthetic carbisocainium chloride in water was investigated by two spectral methods using the probe pyrene. First, the absorption spectroscopy in UV/VIS region was based on studying changes in characteristic absorption spectrum of pyrene in presence of surfactant. The resultant plot of the sum of absorbances for all the major pyrene peaks as a function of the total surfactant concentration shows, around the critical micellar concentration, a typical sigmoidal increase. The fluorescence emission spectroscopy in UV/VIS region of spectrum by the probe pyrene, second procedure, was applied for determination of the cmc from the measurements of the pyrene I1 /I3 ratio as a function of the surfactant concentration. The pyrene ratio data were fitted by the Boltzmann-type sigmoid of decreasing character.

scholarly journals Sequences flanking the transmembrane segments facilitate mitochondrial localization and membrane fusion by mitofusin

2017 ◽  
Vol 114 (46) ◽  
pp. E9863-E9872 ◽  
Author(s):  
Xiaofang Huang ◽  
Xin Zhou ◽  
Xiaoyu Hu ◽  
Amit S. Joshi ◽  
Xiangyang Guo ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Fusion ◽  
Amphipathic Helix ◽  
Mitochondrial Localization ◽  
Mitochondrial Membranes ◽  
Gtpase Domain ◽  
Hydrophobic Residues ◽  
Transmembrane Segments ◽  
Mechanistic Insight ◽  
Insight Into

Mitochondria constantly divide and fuse. Homotypic fusion of the outer mitochondrial membranes requires the mitofusin (MFN) proteins, a family of dynamin-like GTPases. MFNs are anchored in the membrane by transmembrane (TM) segments, exposing both the N-terminal GTPase domain and the C-terminal tail (CT) to the cytosol. This arrangement is very similar to that of the atlastin (ATL) GTPases, which mediate fusion of endoplasmic reticulum (ER) membranes. We engineered various MFN-ATL chimeras to gain mechanistic insight into MFN-mediated fusion. When MFN1 is localized to the ER by TM swapping with ATL1, it functions in the maintenance of ER morphology and fusion. In addition, an amphipathic helix in the CT of MFN1 is exchangeable with that of ATL1 and critical for mitochondrial localization of MFN1. Furthermore, hydrophobic residues N-terminal to the TM segments of MFN1 play a role in membrane targeting but not fusion. Our findings provide important insight into MFN-mediated membrane fusion.

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