High-resolution 1H-NMR studies of self-aggregation of melittin in aqueous solution

Brown, L. R.; Lauterwein, Jürgen; Wüthrich, Kurt

doi:10.1016/0005-2795(80)90034-3

Cited by 161 publications

(158 citation statements)

References 11 publications

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“…Under our experimental conditions (10 mM potassium phosphate, pH 7, 150 mM NaCl, 8 nM-44 M melittin), the melittin in solution was predominantly present in an unstructured, most likely monomeric, form ( Fig. 2C) (23)(24)(25). The results presented here can be interpreted in terms of the two-step model, where pore formation (Fig.…”

Section: Resultsmentioning

confidence: 57%

On the Mechanism of Pore Formation by Melittin

Bogaart

Guzman

Mika

et al. 2008

Journal of Biological Chemistry

180

138

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The mechanism of pore formation of lytic peptides, such as melittin from bee venom, is thought to involve binding to the membrane surface, followed by insertion at threshold levels of bound peptide. We show that in membranes composed of zwitterionic lipids, i.e. phosphatidylcholine, melittin not only forms pores but also inhibits pore formation. We propose that these two modes of action are the result of two competing reactions: direct insertion into the membrane and binding parallel to the membrane surface. The direct insertion of melittin leads to pore formation, whereas the parallel conformation is inactive and prevents other melittin molecules from inserting, hence preventing pore formation.Lytic or antimicrobial peptides are small proteins (12-50 amino acids) that affect cells by disrupting the barrier function of lipid membranes (1). There is much interest in these peptides because of their potential pharmaceutical applications, e.g. as cancer drugs and antibiotics (2). Because of their small size and high stability, they can be obtained in large quantities. Of all lytic peptides, the 26-residue melittin is the best studied to date (for review see Ref.3). Melittin is the major constituent of the venom of the European honeybee Apis mellifera. Melittin has been reported to have anticancer effects and is already being used to treat pain and arthritis in Asia (4).Melittin can bind within milliseconds to lipid membranes and adopts an amphipatic ␣-helical conformation, oriented either parallel or perpendicular to the plane of the membrane. The perpendicular conformation is embedded in the membrane and is needed for pore formation, whereas the parallel conformation is inactive (5-10). These observations led to the proposal of a two-step model for pore formation, where melittin at low concentrations binds parallel to the membrane (Fig. 1A, step 1) and at higher concentrations shifts toward the perpendicular orientation (step 2), causing pore formation (10 -12). The transition from parallel to perpendicular is still poorly understood, especially because the cationic (5ϩ at neutral pH) melittin interacts strongly with the lipid headgroups (partitioning coefficient of 10 4 -10 5 M Ϫ1 for phosphatidylcholine (PC) 2 (12-17)), and the energy of the transition must be very high (3,11). Based on the high affinity of melittin for PC headgroups, it has been widely accepted that the concentration of melittin needed for pore formation is not dependent on the absolute melittin concentration but rather on the ratio of melittin to lipid molecules. However, this important implication has never been directly tested by varying the lipid concentration and determining the fractions of bound and free melittin. In this study we analyzed the lipid concentration dependence of melittin action and studied the reversibility of melittin binding, thereby resolving pore formation from binding events. MATERIALS AND METHODSMelittin was purchased from Genscript (Piscataway, NJ), and lipids were from Avanti Polar Lipids (Albaster, AL). Liposomes ...

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Section: Resultsmentioning

confidence: 57%

On the Mechanism of Pore Formation by Melittin

Bogaart

Guzman

Mika

et al. 2008

Journal of Biological Chemistry

180

138

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“…Nonetheless, calculation of the hydrophobic moment 41,57 revealed that the helical residues have high amphiphilic α-helix potential 29,30 , which include Phe22-Leu32 and Cys53-Ala63, Asn68-Val69, Ser92, Lys107 and Asp116-Leu125 ( Figure 5M). In particular, two regions over Phe22-Leu32 and Asp116-Leu125 have amphiphilicity comparable to mellitin, a honeybee membrane-active toxin 39, [58][59][60] , and the intrinsically-unstructured α-synuclein that triggers Parkinson's disease [61][62][63][64][65] . Both of are unstructured in an aqueous solution, and have a high tendency to aggregate, but spontaneously insert into membranes to form amphiphilic α-helices.…”

Section: Discussionmentioning

confidence: 99%

Resolving the paradox for protein aggregation diseases: a common mechanism for aggregated proteins to initially attack membranes without needing aggregates

et al. 2013

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Paradoxically, aggregation of specific proteins is characteristic of many human diseases and aging, yet aggregates have been found to be unnecessary for initiating pathogenesis. Here we determined the NMR topology and dynamics of a helical mutant in a membrane environment transformed from the 125-residue cytosolic all-β MSP by the ALS-causing P56S mutation. Unexpectedly, despite its low hydrophobicity, the P56S major sperm protein (MSP) domain becomes largely embedded in the membrane environment with high backbone rigidity. Furthermore it is composed of five helices with amphiphilicity comparable to those of the partly-soluble membrane toxin mellitin and α-synuclein causing Parkinson's disease. Consequently, the mechanism underlying this chameleon transformation becomes clear: by disrupting the specific tertiary interaction network stabilizing the native all-β MSP fold to release previously-locked amphiphilic segments, the P56S mutation acts to convert the classic MSP fold into a membrane-active protein that is fundamentally indistinguishable from mellitin and α-synuclein which are disordered in aqueous solution but spontaneously partition into membrane interfaces driven by hydrogen-bond energetics gained from forming α-helix in the membrane environments. As segments with high amphiphilicity exist in all proteins, our study successfully resolves the paradox by deciphering that the proteins with a higher tendency to aggregate have a stronger potential to partition into membranes through the same mechanism as α-synuclein to initially attack membranes to trigger pathogenesis without needing aggregates. This might represent the common first step for various kinds of aggregated proteins to trigger familiar, sporadic and aging diseases. Therefore the homeostasis of aggregated proteins is the central factor responsible for in vivo a variety of human diseases including aging. The number and degree of the membrane attacks by aggregated proteins may act as an endogenous clock to count down the aging process. Consequently, a key approach to fight against them is to develop strategies and agents to maintain or even enhance the functions of the degradation machineries.

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“…activity of many other antimicrobial peptides in a similar fashion. For instance, the bee venom peptide, melittin (69), or the human peptide, LL-37 (70), were reported to form aggregates in aqueous solutions. Both peptides are highly hemolytic, whereas their non-aggregated derivatives display significantly reduced hemolytic activity (70).…”

Section: Discussionmentioning

confidence: 99%

Structural Requirements for Potent Versus Selective Cytotoxicity for Antimicrobial Dermaseptin S4 Derivatives

Kustanovich¹,

Shalev²,

Mikhlin³

et al. 2002

Journal of Biological Chemistry

153

169

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To better understand the structural requirements for selective cytotoxicity of antimicrobial peptides, seven dermaseptin S4 analogs were produced and investigated with respect to molecular organization in solution, binding properties to model phospholipid membranes, and cytotoxic properties. Native dermaseptin S4 displayed high aggregation in solution and high binding affinity. These properties correlated with high cytotoxicity. Yet, potency was progressively limited when facing cells whose plasma membrane was surrounded by increasingly complex barriers. Increasing the positive charge of the native peptide led to partial depolymerization that correlated with higher binding affinity and with virtually non-discriminative high cytotoxicity against all cell types. The C-terminal hydrophobic domain was found responsible for binding to membranes but not for their disruption. Truncations of the C terminus combined with increased positive charge of the Nterminal domain resulted in short peptides having similar binding affinity as the parent compound but displaying selective activity against microbes with reduced toxicity toward human red blood cells. Nuclear magnetic resonance-derived three-dimensional structures of three active derivatives enabled the delineation of a common amphipathic structure with a clear separation of two lobes of positive and negative electrostatic potential surfaces. Whereas the spatial positive electrostatic potential extended considerably beyond the peptide dimensions and was required for potency, selectivity was affected primarily by hydrophobicity. The usefulness of this approach for the design of potent and/or selective cytolytic peptides is discussed herein.

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High-resolution 1H-NMR studies of self-aggregation of melittin in aqueous solution

Cited by 161 publications

References 11 publications

On the Mechanism of Pore Formation by Melittin

On the Mechanism of Pore Formation by Melittin

Resolving the paradox for protein aggregation diseases: a common mechanism for aggregated proteins to initially attack membranes without needing aggregates

Structural Requirements for Potent Versus Selective Cytotoxicity for Antimicrobial Dermaseptin S4 Derivatives

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