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1.
This paper describes a convenient oxidative refolding method that exploits the dual property of dimethylsulfoxide (DMSO) as an oxidant and a solvent 'chaperon' in assisting disulfide formation in peptides. Synthetic peptides with preferred secondary structures were used as models. For -sheet peptides, an active fragment of fibroblast growth factor containing the tetrapeptide Arg-Ser-Arg-Arg at a reverse turn was used. A disulfide scan consisting of 16 analogs is designed in which different pairs of amino acids on the antiparallel -sheets flanking the active determinants are replaced with cysteine. DMSO in aqueous buffer at pH 6 or 8 was found to minimize aggregation due to -sheet formation in all 16 -stranded peptides and provided monocyclic disulfide peptides within 7 h. In contrast, air oxidation required a significantly longer duration for completion under similar conditions, and only 8 of 16 peptides formed intramolecular disulfides. Rate accelerations at pH 8 were found in exo-disulfide formation involving peptides with amino terminal cysteine, irrespective of oxidation conditions. The exo-disulfide effect in accelerating disulfide formation may be useful for regiospecific disulfide formation. For -helix peptides, the two-disulfide endothelin (ET) was used as a model. DMSO in combination with trifluoroethanol (TFE) was found to favor the desired bicyclic 1,4-disulfide bridged ET (1,4-ET) over the incorrectly folded 1,3-ET. Under aqueous conditions at pH 5–11, 1,4-ET to 1,3-ET was formed in the ratio of approximately 3:1, while the use of DMSO and TFE increased the ratio to 11:1. This solvent combination may stabilize an -helical stretch found in ET and contribute to enhanced selectivity. Thus, our results show that DMSO in disulfide formation in an aqueous or helix-promoting solution may serve as an oxidant and a 'chaperon' solvent system to provide regiospecificity for oxidative disulfide formation. 相似文献
2.
Human Antimicrobial Peptides: Defensins, Cathelicidins and Histatins 总被引:12,自引:0,他引:12
Antimicrobial peptides, which have been isolated from many bacteria, fungi, plants, invertebrates and vertebrates, are an
important component of the natural defenses of most living organisms. The isolated peptides are very heterogeneous in length,
sequence and structure, but most of them are small, cationic and amphipathic. These peptides exhibit broad-spectrum activity
against Gram-positive and Gram-negative bacteria, yeasts, fungi and enveloped viruses. A wide variety of human proteins and
peptides also have antimicrobial activity and play important roles in innate immunity. In this review we discuss three important
groups of human antimicrobial peptides. The defensins are cationic non-glycosylated peptides containing six cysteine residues
that form three intramolecular disulfide bridges, resulting in a triple-stranded β-sheet structure. In humans, two classes
of defensins can be found: α-defensins and β-defensins. The defensin-related HE2 isoforms will also be discussed. The second
group is the family of histatins, which are small, cationic, histidine-rich peptides present in human saliva. Histatins adopt
a random coil conformation in aqueous solvents and form α-helices in non-aqueous solvents. The third group comprises only
one antimicrobial peptide, the cathelicidin LL−37. This peptide is derived proteolytically from the C-terminal end of the human CAP18 protein. Just like the histatins, it adopts a largely random coil conformation in a hydrophilic
environment, and forms an α-helical structure in a hydrophobic environment. 相似文献
3.
Christine Talmard Rodrigue Leuma Yona Peter Faller 《Journal of biological inorganic chemistry》2009,14(3):449-455
The amyloidoses are a group of disorders characterized by aberrant protein folding and assembly, leading to the deposition
of insoluble protein fibrils (amyloid), which provokes cell dysfunction and later cell death. One of the physiologically relevant
environmental factors able to affect the conformation and hence the aggregation properties of amyloidogenic proteins/peptides
is metal ions. Zn(II) promotes aggregation of most amyloidogenic peptides/proteins in vitro, including amyloid β protein (Aβ),
but the underlying mechanism is not known. To better understand this mechanism the present study focused on the partially
α-helical conformer, supposed to be an intermediate in Aβ aggregation. This partially α-helical conformer is stabilized by
10–20% 2,2,2-trifluoroethanol (TFE): therefore, the influence of Zn binding on the aggregation of the amylidogenic model peptide
Aβ(1–28) (Aβ28) was investigated at different TFE concentrations. The results showed a synergistic effect of Zn(II) and 10%
TFE, i.e., that either Zn or 10% TFE accelerated Aβ28 aggregation on its own, but with them together an at least 10 times
promotion of Aβ28 aggregation was observed. Further studies by thioflavin T fluorescence spectroscopy, transmission electron
microscopy, and circular dichroism (CD) spectroscopy suggested that the aggregates of Zn-Aβ28 formed in 10%TFE contain a β-sheet
secondary structure and are more of the amyloid type. CD spectroscopy indicated that Zn binding disrupted partially the α-helical
structure of Aβ28 in TFE. Thus, we propose that the promotion of Aβ28 aggregation by Zn is based on the transformation of
the partially α-helical conformer (intermediate) towards the β-sheet amyloid structure by a destabilization of the α-helix
in the intermediate.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.
相似文献
Peter FallerEmail: Email: |
4.
B. Ahmad S. K. Haq A. Varshney A. A. Moosavi-Movahedi R. H. Khan 《Biochemistry. Biokhimii?a》2010,75(4):486-493
We have studied the effect of trifluoroethanol (TFE) on the native (pH 7.0), acid unfolded (pH 2.6), and molten globule (pH
1.4) states of glucose oxidase (GOX) by circular dichroism and fluorescence spectroscopy. In the presence of 50% TFE, at pH
7.0 and 2.6, GOX exhibited a transition from native coiled-coil and acid unfolded state to non-associating α-helical state.
Interestingly, at pH 1.4, 15% TFE induced the formation of β-structured intermediate by loss of 1-anilino-8-naphthalenesulfonate
binding site and almost all tertiary contacts. The β-structured intermediate converted into open helical conformation on further
addition of TFE. 相似文献
5.
Heitz Annie Le-Nguyen Dung Castro Bertrand Chiche Laurent 《International journal of peptide research and therapeutics》1997,4(4-6):245-249
Summary Trypsin inhibitor EETI II, possessing six cysteines engaged in three disulfide bridges, shares a common structural motif with
other proteins of different origins and functions. To understand the principles that govern folding of this largely distributed
basic scaffold, mainly composed of a small triple-stranded β-sheet, we have studied different stages in the folding of EETI
II. The conformational properties of a synthetic analogue of EETI II possessing only one native (15–27) disulfide bridge were
investigated with the combined use of1H NMR and molecular modelling. Although two native-like reverse turns were observed, formation of β-sheet could not be evidenced
in the one disulfide analogue, while the motif has been shown to be present in a folding intermediate with two native disulfide
bridges (9–21 and 15–27). These results suggest that the structural motif requires stabilisation by two disulfide bridges. 相似文献
6.
Sergei Andreev Igor Andreev Elena Nikolaeva Anna Petrukhina Vladimir Zemskov Mariam Vafina 《International journal of peptide research and therapeutics》1998,5(2-3):63-66
Summary The V3 loop from HIV-1 envelope glycoprotein gp120 is involved in viral entry and determines the cellular tropism and HIV-1-induced
cell-cell fusion. Earlier we have shown that V3 loop peptides representing the sequences of syncytia-inducing HIV strains
have high membranotropic activity. These peptides caused the lysis of liposomes of various lipid compositions, could fuse
negatively charged liposomes and induced hemolysis of erythrocytes. In contrast, peptides mimicking the sequences of non-syncytia-inducing
viruses showed no lytic or fusion activities at the same concentrations. Now we have found that the V3 loop synthetic peptides
containing the conserved GPGR region, derived from T-lymphotropic strains (BRU and MN), as opposed to peptides containing
the GPGQ region, are able to cause a pronounced membrane permeabilization (dissipation of the ΔpH and the Δψ) of human peripheral
blood lymphocytes, erythrocytes and plasma membrane vesicles at micromolar concentrations with a dose-dependent kinetics.
Analysis of the secondary structures of the peptides by circular dichroism revealed conformational changes in V3 loop peptides
depending on solvent hydrophobicity: from random coil in water to an α-helix/β-sheet conformation in trifluoroethanol. Such
structural changes of the V3 loop together with the membrane insertion of the gp41 N-terminal fusion peptide may promote the
formation of the fusion pore during virus-cell fusion. 相似文献
7.
Emilia Bramanti Francesco Lenci Antonella Sgarbossa 《European biophysics journal : EBJ》2010,39(11):1493-1501
We have determined the secondary structure of 1–40 β-amyloid peptides by Fourier-transform infrared spectroscopy (FTIR) and
characterized the peptide photophysical properties before and after self-assembly by using intrinsic tyrosine steady-state
and time-resolved fluorescence. All measurements were performed in the presence and absence of hypericin (Hyp), an exogenous
natural polycyclic pigment that has been shown to inhibit fibril formation and has also been used as a fluorescent probe.
We monitored the time course of the aggregation process measuring 405 nm light diffusion at 90° and used thioflavin T to reveal
the presence of fibrils. FTIR quantitative analysis evidenced a prevalent random conformation at t = 0 with and without Hyp. Fibrils showed a predominant parallel β-sheet structure and a small percentage of α-helix. The
results of fluorescence measurements showed that Hyp does significantly interact with peptides in β-sheet conformation. In
conclusion, hypericin does hinder the formation of fibrils, but the percentages of parallel β-sheets were not significantly
different from those found in samples not treated with Hyp. 相似文献
8.
Ollesch J Poschner BC Nikolaus J Hofmann MW Herrmann A Gerwert K Langosch D 《European biophysics journal : EBJ》2008,37(4):435-445
LV-peptides were designed as membrane-spanning low-complexity model structures that mimic fusion protein transmembrane domains.
These peptides harbor a hydrophobic core sequence that consists of helix-promoting and helix-destabilizing residues at different
ratios. Previously, the fusogenicity of these peptides has been shown to increase with the conformational flexibility of their
hydrophobic cores as determined in isotropic solution. Here, we examined the secondary structure, orientation, and distribution
of LV-peptides in membranes. Our results reveal that the peptides are homogeneously distributed within the membranes of giant
unilamellar liposomes and capable of fusing them. Increasing the valine content of the core up to the level of the β-branched
residue content of SNARE TMDs (∼50%) enhances fusogenicity while maintaining a largely α-helical structure in liposomal membranes.
A further increase in valine content or introduction of a glycine/proline pair favors β-sheet formation. In planar bilayers,
the α-helices adopt oblique angles relative to the bilayer normal and the ratio of α-helix to β-sheet responds more sensitively
to valine content. We propose that the fusogenic conformation of LV-peptides is likely to correspond to a membrane-spanning
α-helix. β-Sheet formation in membranes may be considered a side-reaction whose extent reflects conformational flexibility
of the core. 相似文献
9.
We have studied the effect of 2,2,2-trifluoroethanol (TFE), an α-helix inducer, versus methyl cyanide (MeCN), a β-sheet inducer,
on acid-denatured human serum albumin (HSA) using far-UV circular dichroism, intrinsic fluorescence, 1-anilino-8-naphthalene
sulfonate binding, and acrylamide quenching studies. Interestingly, at pH 2.0, where the recovery and resolution of the protein
in reverse phase chromatography is high, its secondary structure remains unchanged even in the presence of very high concentration
(76% v/v) of MeCN. Gain of 23 and 34% α-helicity was observed in the presence of 20 and 50% TFE, respectively. At pH 7.3,
HSA aggregates in the presence of 40% MeCN, but it remains soluble up to 75% MeCN at pH 2.0. The results seem to be important
for HSA isolation and purification. 相似文献
10.
Shah V Baldrian P Eichlerova I Dave R Madamwar D Nerud F Gross R 《Biotechnology letters》2006,28(9):651-655
Dimethyl sulfoxide (DMSO) is commonly used as a co-solvent to dissolve poorly water-soluble biologically active agents to
assess their biological activities such as for enzyme induction. The question addressed was whether DMSO can be assumed to
be an inert co-solvent. The influence of DMSO on the production of extracellular enzymes by Pleurotus ostreatus was investigated. DMSO functioned as either an inducer or a repressor, depending on the enzyme studied. The production of
laccase and endo-1,4-β-xylanase increased by 29 and 250%, respectively, in presence of DMSO. However, DMSO repressed the activities of manganese
peroxidase, β-glucosidase, β-xylanase, and endo-1,4-β-glucanase by 30, 33, 99 and 16%, respectively. These results raise concerns about the interpretation of bioactivity measurements
when DMSO is assumed to function as an inert co-solvent to solubilize water-insoluble molecules.
Revisions requested 20 December 2005; Revisions received 6 February 2006 相似文献
11.
Fluorinated alcohols such as hexafluoroisopropanol (HFIP) and trifluoroethanol (TFE) have the ability to promote α-helix and β-hairpin structure in proteins and peptides. HFIP has been used extensively to dissolve various amyloidogenic proteins and peptides including Aβ, in order to ensure their monomeric status. In this paper, we have investigated the self-assembly of Aβ40, Aβ42, and Aβ43 in aqueous mixtures of fluorinated alcohols from freshly dissolved stock solutions in HFIP. We have observed that formation of fibrillar and non-fibrillar structures are dependent on the solvent composition. Peptides form fibrils with ease when reconstituted in deionized water from freshly dissolved HFIP stocks. In aqueous mixtures of fluorinated alcohols, either predominant fibrillar structures or clustered aggregates were observed. Aqueous mixtures of 20% HFIP are more favourable for Aβ fibril formation as compared to 20% TFE. When Aβ40, Aβ42, and Aβ43 stocks in HFIP are diluted in 50% aqueous mixtures in phosphate buffer or deionized water followed by slow evaporation of HFIP, Aβ peptides form fibrils in phosphate buffer and deionized water. The clustered structures could be off-pathway aggregates. Aβ40, Aβ42, and Aβ43 showed significant α-helical content in freshly dissolved HFIP stocks. The α-helical conformational intermediate in Aβ40, Aβ42, and Aβ43 could favour the formation of both fibrillar and non-fibrillar aggregates depending on solvent conditions and rate of α-helical to β-sheet transition. 相似文献
12.
Native human β-casein (CN) at all phosphorylation levels exhibits reproducible behavior and appears to have a unique, stable
folding pattern. In contrast, the recombinant non-phosphorylated form of human β-CN (β-CN-0P) with the exact amino acid sequence
(wild-type), expressed and purified from Escherichia coli, differs greatly in its behavior from the native protein and the complexes formed are unstable to thermal cycling. However,
when it was incorporated into reconstituted milk micelles, using bovine κ-CN at a κ/β molar ratio of 1/3 with added Ca2+ ions and inorganic phosphate (Pi) at levels that would ordinarily precipitate, its association behavior vs. temperature as monitored by turbidity (OD400 nm) approximated that of native β-CN-0P. This suggests that the milk micelle system, and particularly the colloidal calcium
phosphate, may act as a ‘molecular chaperon’ to direct the folding of the molecule into the highly stable conformation found
in the purified native human β-CN molecule. 相似文献
13.
The main component of senile plaques found in AD brain is amyloid β-peptide (Aβ), and the neurotoxicity and aggregation of
Aβ are associated with the formation of β-sheet structure. Experimentally, beta sheet breaker (BSB) peptide fragment Leu-Pro-Phe-Phe-Asp
(LPFFD) can combine with Aβ, which can inhibit the aggregation of Aβ. In order to explore why LPFFD can inhibit the formation
of β-sheet conformation of Aβ at atomic level, first, molecular docking is performed to obtain the binding sites of LPFFD
on the Aβ(1–42) (LPFFD/Aβ(1–42)), which is taken as the initial conformation for MD simulations. Then, MD simulations on LPFFD/Aβ(1–42)
in water are carried out. The results demonstrate that LPFFD can inhibit the conformational transition from α-helix to β-sheet
structure for the C-terminus of Aβ(1–42), which may be attributed to the hydrophobicity decreasing of C-terminus residues
of Aβ(1–42) and formation probability decreasing of the salt bridge Asp23-Lys28 in the presence of LPFFD. 相似文献
14.
Afonso M. S. Duarte Edwin R. de Jong Rob B. M. Koehorst Marcus A. Hemminga 《European biophysics journal : EBJ》2010,39(4):639-646
The conformation of a transmembrane peptide, sMTM7, encompassing the cytoplasmic hemi-channel domain of the seventh transmembrane
section of subunit a from V-ATPase from Saccharomyces cerevisiae solubilized in SDS solutions was studied by circular dichroism (CD) spectroscopy and fluorescence spectroscopy of the single
tryptophan residue of this peptide. The results show that the peptide adopts an α-helical conformation or aggregated β-sheet
depending on the peptide-to-SDS ratio used. The results are compared with published data about a longer version of the peptide
(i.e., MTM7). It is concluded that the bulky, positively charged arginine residue located in the center of both peptides has
a destabilizing effect on the helical conformation of the SDS-solubilized peptides, leading to β-sheet formation and subsequent
aggregation. 相似文献
15.
Vidal Pierre Morris May C. Chaloin Laurent Méry Jean Heitz Frédéric Divita Gilles 《International journal of peptide research and therapeutics》1997,4(4-6):227-230
Summary We describe the synthesis of an amphipathic vector peptide which is able to form complexes with nucleic acids. Based on circular
dichroism investigations, the nature of the structure obtained in water is questioned. The peptide adopts an α-helical structure
in TFE and is partially in a β-sheet conformation in phosphate buffer at low peptide concentrations. 相似文献
16.
Powerful solvent systems useful for synthesis of sparingly-soluble peptides in solution. 总被引:3,自引:0,他引:3
H Kuroda Y N Chen T Kimura S Sakakibara 《International journal of peptide and protein research》1992,40(3-4):294-299
Our maximum protection strategy for the synthesis of human parathyroid hormone(1-84) indicates that fully protected peptide segments in the form of Boc-peptide phenacyl (Pac) ester are relatively soluble in ordinary organic solvents such as DMF, NMP or DMSO, which are suitable for coupling segments. However, about 1% of such segments synthesized were found to be insoluble even in the most polar solvent, DMSO. Thus, a more powerful solvent which can be used for their peptide synthesis was pursued. Among the solvent systems tested, a mixture of trifluoroethanol (TFE) or hexafluoroisopropanol (HFIP) and trichloromethane (TCM) or dichloromethane (DCM) was found to be most powerful for dissolving such sparingly-soluble protected peptides. These solvent systems were confirmed to be useful for the removal reaction of the carboxy-terminal Pac esters from the sparingly-soluble segments. They were then tested for the coupling reactions of fully protected Boc-peptides with other sparingly-soluble peptide esters. The TFE/TCM or TFE/DCM system was extremely useful for coupling segments without danger of racemization and of trifluoroester formation, if WSCI was used as the coupling reagent in the presence of 3,4-dihydro-3-hydroxy-4-oxo-1,2,3-benzotriazine (HOOBt). 相似文献
17.
Tengku Haziyamin Tengku Abdul Hamid Raja Noor Zaliha Raja Abd Rahman Abu Bakar Salleh Mahiran Basri 《The protein journal》2010,29(4):290-297
The use of lipase in hydrophilic solvent is usually hampered by inactivation. The solvent stability of a recombinant solvent
stable lipase isolated from thermostable Bacillus sp. strain 42 (Lip 42), in DMSO and methanol were studied at different solvent-water compositions. The enzymatic activities
were retained in up to 45% v/v solvent compositions. The near-UV CD spectra indicated that tertiary structures were perturbed
at 60% v/v and above. Far-UV CD in methanol indicated the secondary structure in Lip 42 was retained throughout all solvent
compositions. Fluorescence studies indicated formations of molten globules in solvent compositions of 60% v/v and above. The
enzyme was able to retain its secondary structures in the presence of methanol; however, there was a general reduction in
β-sheet and an increase in α-helix contents. The H-bonding arrangements triggered in methanol and DMSO, respectively, caused
different forms of tertiary structure perturbations on Lip 42, despite both showing partial denaturation with molten globule
formations. 相似文献
18.
Clippingdale Andrew B. He Wei-Lan Macris Mary Wade John D. Barrow Colin J. 《International journal of peptide research and therapeutics》1999,6(5-6):289-293
Summary Modifications to secondary structure and fibril formation caused by multiple acetylHmb backbone amide protection of Alzheimer's
disease Aβ(1–40) were investigated using circular dichroism spectroscopy and electron microscopy. Penta(acetylHmb) Aβ(1–40)
was observed to have a reduced ability to form α-helix and β-sheet structures under the same solution conditions as the native
peptide, with α-helical propensity being reduced more significantly than β-sheet propensity. Further, acetylHmb backbone protection
was found to alter Aβ(1–40) interaction with SDS-micelles by preventing α-helix formation. Aβ fibril formation, a characteristic
property of this peptide, was also not observed for penta(acetylHmb) Aβ(1–40). 相似文献
19.
Bourbo V Matmor M Shtelman E Rubinov B Ashkenasy N Ashkenasy G 《Origins of life and evolution of the biosphere》2011,41(6):563-567
Most self-replicating peptide systems are made of α-helix forming sequences. However, it has been postulated that shorter
and simpler peptides may also serve as templates for replication when arranged into well-defined structures. We describe here
the design and characterization of new peptides that form soluble β-sheet aggregates that serve to significantly accelerate
their ligation and self-replication. We then discuss the relevance of these phenomena to early molecular evolution, in light
of additional functionality associated with β-sheet assemblies. 相似文献
20.
D. Walker P. Ledesma O.D. Delgado J.D. Breccia 《World journal of microbiology & biotechnology》2006,22(7):761-764
Summary The strain SES28 was isolated from an indoor contaminated agar plate during a screening program for alkaliphilic CM-cellulose-degrading
bacteria. It showed a prominent clear hydrolysis of the substrate at pH 10. The 16S rDNA analysis related it to the genus
Nocardiopsis
.
Nocardiopsis sp. SES28 was able to grow at pH values up to 10.5, the major biomass being produced at pH 10, and pH 8 was the optimum for
β-1,4-glucanase production. The optimum pH for β-1,4-glucanase activity was 9.0, and it was higher than 60% throughout the
pH range 6.5–10.0; showing 94% of its a relative activity at pH 10. The feature of this bacterium to produce β-1,4-glucanase
active in a broad pH-range might be useful for detergent- and textile-processing technologies. 相似文献