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The lamellar gel/liquid-crystalline and the lamellar liquid-crystalline/reversed hexagonal phase transitions of aqueous dispersions of a number of synthetic phosphatidylethanolamines containing linear saturated, branched chain, and alicyclic fatty acyl chains of varying length were studied by differential scanning calorimetry, 31P nuclear magnetic resonance spectroscopy, and X-ray diffraction. For any given homologous series of phosphatidylethanolamines containing a single chemical class of fatty acids, the lamellar gel/liquid-crystalline phase transition temperature increases and the lamellar liquid-crystalline/reversed hexagonal phase transition temperature decreases with increases in hydrocarbon chain length. For a series of phosphatidylethanolamines of the same hydrocarbon chain length but with different chemical structures, both the lamellar gel/liquid-crystalline and the lamellar liquid-crystalline/reversed hexagonal phase transition temperatures vary markedly and in the same direction. In particular, at comparable effective hydrocarbon chain lengths, both the lamellar gel/liquid-crystalline and the lamellar liquid-crystalline/reversed hexagonal phase transition temperatures vary in parallel, such that the temperature difference between these two phase transitions is nearly constant. Moreover, at comparable effective acyl chain lengths, the d spacings of the lamellar liquid-crystalline phases and of the inverted hexagonal phases are all similar, implying that the thickness of the phosphatidylethanolamine bilayers at the onset of the lamellar liquid-crystalline/reversed hexagonal phase transition and the diameter of the water-filled cylinders formed at the completion of this phase transition are comparable and independent of the chemical structure of the acyl chain.(ABSTRACT TRUNCATED AT 250 WORDS)

Growth and membrane lipid properties of Acholeplasma laidlawii B lacking fatty acid heterogeneity

The physical state of the membrane lipids, as determined by fatty acid composition and environmental temperature, has a marked effect on both the temperature range within which Acholeplasma laidlawii B cells can grow and on growth rates within the permissible temperature ranges. The minimum growth temperature of 8 °C is not defined by the fatty acid composition of the membrane lipids when cells are enriched in fatty acids giving rise to gel to liquid-crystalline membrane lipid phase transitions occurring below this temperature. The elevated minimum growth temperatures of cells enriched in fatty acids giving rise to lipid phase transitions occurring at higher temperatures, however, are clearly defined by the fatty acid composition of the membrane lipids. The optimum and maximum growth temperatures are also influenced indirectly by the physical state of the membrane lipids, being significantly reduced for cells supplemented with lower melting, unsaturated fatty acids. The temperature coefficient of growth at temperatures near or above the midpoint of the lipid phase transition is 16 to 18 kcal mol , but this value increases abruptly to 40 to 45 kcal mol at temperatures below the phase transition midpoint. Both the absolute rates and temperature coefficients of cell growth are similar for cells whose membrane lipids exist entirely or predominantly in the liquid-crystalline state, but absolute growth rates decline rapidly and temperature coefficients increase at temperatures where more than half of the membrane lipids become solidified. Cell growth ceases when the conversion of the membrane lipid to the gel state approaches completion, but growth and replication can continue at temperatures where less than one tenth of the total lipid remains in the fluid state. An appreciable heterogeneity in the physical state of the membrane lipids can apparently be tolerated by this organism without a detectable loss of membrane function.

Many of the contributions in this volume are concerned with the physical measurement and biochemical regulation of membrane lipid fluidity in prokaryotic microorganisms. This contribution is primarily concerned with the biological function of such regulatory mechanisms and with the physiological consequences of their absence or impairment. In the first part of this essay, some brief comments on the concept of fluidity as applied to membranes are offered and a short, critical analysis of the physical techniques available for the measurement of lipid fluidity and phase state is given. Next, a summary and critical discussion of what we currently know about the relationship between membrane lipid fluidity and phase state and the ability of bacteria and mycoplasmas to grow at various temperatures are presented. Related work on the influence of the fatty acid composition of the lipids of bacterial membranes on their ability to survive exposure to extremes of temperature is then reviewed. Finally, the possible molecular bases for the observed relationships between membrane lipid fluidity and phase state and the growth and survival of prokaryotic microorganisms are discussed, as is the biological significance of homeoviscous or homeophasic regulatory mechanisms in bacteria and mycoplasmas.

The extent of incorporation of a wide variety of exogenous saturated, unsaturated, branched-chain, and cyclopropane fatty acids into the membrane lipids of Acholeplasma laidlawii B was systematically studied. Within each fatty acid class the extent of incorporation generally increased markedly with increasing chain length, reached a maximum, and then declined progressively but less sharply with further increases above that chain length giving maximal direct incorporation. Certain shorter-chain members of each fatty acid class underwent complete or partial conversion to longer-chain homologues before utilization for complex lipid biosynthesis. The degree and extent of chain elongation and direct incorporation and the characteristic dependence of each of these processes on fatty acid chain length and structure correlated well with the physical properties (melting temperatures) of the exogenous fatty acids. The in vivo specificity of the enzyme systems responsible for the incorporation of exogenous fatty acids was such that the fluidity and physical state of the membrane lipids were maintained within a definite, albeit a relatively wide, range. We also observed that the neutral glycolipids typically have similar fatty acid compositions, which are somewhat different from those of the major phosphatides, which also exhibit similar fatty acid spectra. The phosphorylated glycolipid glycerophosphoryldiglucosyl diglyceride, however, always maintained a unique fatty acid composition quite different from that of the diglucosyl diglyceride from which it is presumably derived. These characteristic differences in fatty acid composition appear to function to minimize differences in phase transition temperatures, thus producing a more physicochemically homogeneous mixture of membrane lipids than would result from a nonspecific incorporation of fatty acids.

The thermotropic phase behavior of aqueous dispersions of phosphatidylcholines containing one of a series of methyl iso-branched fatty acyl chains was studied by differential scanning calorimetry. These compounds exhibit a complex phase behavior on heating which includes two endothermic events, a gel/gel transition, involving a molecular packing rearrangement between two gel-state forms, and a gel/liquid-crystalline phase transition, involving the melting of the hydrocarbon chains. The gel to liquid-crystalline transition is a relatively fast, highly cooperative process which exhibits a lower transition temperature and enthalpy than do the chain-melting transitions of saturated straight-chain phosphatidylcholines of similar acyl chain length. In addition, the gel to liquid-crystalline phase transition temperature is relatively insensitive to the composition of the aqueous phase. In contrast, the gel/gel transition is a slow process of lower cooperativity than the gel/liquid-crystalline phase transition and is sensitive to the composition of the bulk aqueous phase. The gel/gel transitions of the methyl iso-branched phosphatidylcholines have very different thermodynamic properties and depend in a different way on hydrocarbon chain length than do either the "subtransitions" or the "pretransitions" observed with linear saturated phosphatidylcholines. The gel/gel and gel/liquid-crystalline transitions are apparently concomitant for the shorter chain iso-branched phosphatidylcholines but diverge on the temperature scale with increasing chain length, with a pronounced odd/even alternation of the characteristic temperatures of the gel/gel transition.(ABSTRACT TRUNCATED AT 250 WORDS)

The physical state of the membrane lipids, as determined by fatty acid composition and environmental temperature, has a marked effect both on the temperature range within which A. laidlawii can grow and on the temperature coefficient of growth within the permissible temperature range. The minimum growth temperature under certain conditions is clearly defined by the lower boundary of the gel–to–liquid-crystalline phase transition of the membrane lipids. The physical state of the membrane lipids can also influence the optimum and maximum growth temperatures. An a brupt increase in the temperature coefficient of growth is noted at temperatures between the phase transition boundaries. Both the absolute rates and the temperature coefficients of cell growth are similar for cells whose membrane lipids exist entirely or predominantly in the liquid-crystalline state, but absolute growth rates decline rapidly and temperature coefficients increase when most of the membrane lipids become solidified. Some cell growth, however, can continue at temperatures at which less than 10% of the total lipid remains in the fluid state. Conversion of the membrane lipid from the liquid-crystalline to the gel state is accompanied by a progressive aggregation of intramembranous protein particles. An appreciable heterogeneity in the physical state of the membrane lipids can apparently be tolerated by this organism without a detectable loss of membrane function.

The thermotropic phase behavior of aqueous dispersions of nine dl-methyl branched anteisoacylphosphatidylcholines was studied by differential scanning calorimetry and /sup 31/P nuclear magnetic resonance spectroscopy. The calorimetric studies demonstrate that these compounds all exhibit a complex phase behavior, consisting of at least two minor, low-enthalpy, gel-state transitions which occur at temperatures just prior to the onset of the gel/liquid-crystalline phase transition. In addition, at still lower temperatures, anteisobranched phosphatidylcholines containing fatty acyl chains with an odd number of carbon atoms show a major, higher enthalpy, gel-state transition, which was assigned to a conversion from a condensed to a more loosely packed gel phase. /sup 31/P NMR spectroscopic studies indicate that the major gel-state transition is accompanied by a considerable change in the mobility of the phosphate head group and that, at temperatures just prior to the onset of the gel/liquid-crystalline phase transition, the mobility of the phosphate head group is comparable to that normally exhibited by the liquid-crystalline state of most other phospholipids. The temperatures at which the gel/liquid-crystalline phase transition occurs and the enthalpy change associated with this process are considerably lower than those of the saturated n-acyl-PC's of comparable acyl chain length. The calorimetric and /sup 31/P nuclearmore » magnetic resonance data thus indicate that, although the methyl anteisobranched phosphatidylcholines can form a fairly condensed gel state at sufficiently low temperatures, these phospholipids normally adopt a more loosely packed gel state, and possibly a more ordered liquid-crystalline state, than do phosphatidylcholines containing linear saturated fatty acids of comparable chain length.« less

The thermotropic phase behavior of aqueous dispersions of 10 phosphatidylcholines containing omega-cyclohexyl-substituted acyl chains was studied by differential scanning calorimetry and 31P nuclear magnetic resonance spectroscopy. The presence of the omega-cyclohexyl group has a profound effect on the thermotropic phase behavior of these compounds in a manner dependent on whether the fatty acyl chains have odd- or even-numbered linear carbon segments. The thermotropic phase behavior of the odd-numbered phosphatidylcholines is characterized by a single heating endotherm that was shown to be a superposition of at least two structural events by calorimetric cooling experiments. 31P NMR spectroscopy also showed that the single endotherm of the odd-chain compounds is the structural equivalent of a concomitant gel-gel and gel to liquid-crystalline phase transition. The calorimetric behavior of the even-numbered phosphatidylcholines is characterized by a complex array of gel-state phenomena, in addition to the chain-melting transition, in both the heating and cooling modes. The gel states of these even-numbered compounds are characterized by a relatively greater mobility of the phosphate head group as seen by 31P NMR spectroscopy. The differences between the odd-numbered and even-numbered compounds are reflected in a pronounced odd-even alternation in the characteristic transition temperatures and enthalpies and in differences in their responses to changes in the composition of the bulk aqueous phase. Moreover, both the odd-numbered and even-numbered omega-cyclohexylphosphatidylcholines exhibit significantly lower chain-melting transition temperatures and enthalpies than do linear saturated phosphatidylcholines of comparable chain length.(ABSTRACT TRUNCATED AT 250 WORDS)

While adaptation of microbial membrane lipid composition is widely observed in response to changes in temperature, and in some cases to the presence of membrane-fluidizing substances, the physiological importance of such adaptations remains largely unexplored. To shed some light on this question question we have used our ability to extensively vary the membrane fatty acid composition of Acholeplasma laidalawii B to study a variety of membrane functions is membranes whose lipids contain a single fatty acyl species whose structure and physical properties can be widely varried. Our results suggest that control of the membrane lipid phase state is considerably more important to the proper functioning of the organism than is control of the ‘fluidity’ of liquid-crystalline lipids.

The positional distributions of a series of fifteen positional isomers of cis-octadecenoic acid in the phosphatidylglycerol synthesized by Acholeplasma laidlawii B were determined. The specificity for the 2-position generally increased (and that for the 1-position decreased) as the position of the double bond moved from the methyl terminus toward the carbonyl group of the hydrocarbon chain. The Δ5 and Δ6cis-octadecenoates, however, were exceptional in exhibiting a much lower 2-positional specificity than the adjacent members in this series. The positional specificities of the Δ5, Δ6 and Δ9 through Δ17-cis-octadecenoic acids correlated well with the melting behavior (gel to liquid-crystalline phase transition temperatures) of the corresponding 1,2-dioctadecenoyl phosphatidylcholines. In contrast, the Δ3 and Δ4, and the Δ7 and Δ8-octadecenoate isomers exhibited specificities for position 2 which are markedly and moderately greater, respectively, than would be predicted from the melting behavior of the corresponding phosphatidylcholines. The fatty acid specificity of the enzyme system catalyzing the acylation of sn-glycerol-3-phosphate in this organism is such that the biosynthesis of membrane glycerolipids having a high degree of physicochemical homogeneity is generally promoted, particularly when naturally occurring mono- and polyunsaturated fatty acids are utilized.

Publisher Summary The fluidity and physical state of the membrane lipids affect the activity of membrane enzymes and transport systems. Proteins are free to diffuse laterally in the plane of the membrane unless constrained by their interactions with other proteins, either within the membrane itself or outside the membrane proper, including proteins functioning as cytoskeletal structural elements. The differential lateral mobility of proteins within the lipid bilayer leads to a locally heterogeneous or mosaic two- dimensional organization of the membrane. This chapter focuses on studies of intact microbial membranes that are associated with the effect of the physical properties of membrane lipids on the functioning of particular membrane enzymes and transport systems. Complex physiological functions, including cell growth are also discussed. In addition, the chapter summarizes the effects of lipid on the activities of isolated membrane-associated enzymes. Properties of membrane constituents (such as membrane lipids and membrane proteins) are discussed; and the chapter presents an account of Arrhenius plots of membrane transport systems and enzymes. Studies of isolated and lipid-reconstituted membrane enzymes and transport proteins have demonstrated that lipids can function to solubilize water-insoluble substrates or products, to solubilize and disperse the enzyme or transport protein, and to provide a proper environment for stabilizing the protein of interest in an active conformation. Both the fluidity and interfacial properties of the lipids employed in reconstitution studies can be important in determining the activity of membrane proteins and the regulation of this activity by ligand binding. However, until the nature of lipid–protein interactions at the molecular level is understood, and until the secondary and tertiary structures of integral membrane proteins in a lipid milieu are determined, the molecular mechanisms that help the lipids to modulate the function of membrane proteins will remain obscure.

The active transport of sodium ions in live Acholeplasma laidlawii B cells and in lipid vesicles containing the (Na+-Mg2+)-ATPase from the plasma membrane of this microorganism was studied by 23Na nuclear magnetic resonance spectroscopic and 22Na tracer techniques, respectively. In live A. laidlawii B cells, the transport of sodium was an active process in which metabolic energy was harnessed for the extrusion of sodium ions against a concentration gradient. The process was inhibited by low temperatures and by the formation of gel state lipid in the plasma membrane of this organism. In reconstituted proteoliposomes containing the purified (Na+-Mg2+)-ATPase, the hydrolysis of ATP was accompanied by the transport of sodium ions into the lipid vesicles, and the transport process was impaired by reagents known to inhibit ATPase activity. At the normal growth temperature (37 degrees C), this transport process required a maximum of 1 mol of ATP per mol of sodium ion transported. Together, these results provide direct experimental evidence that the (Na+-Mg2+)-ATPase of the Acholeplasma laidlawii B membrane is the cation pump which maintains the low levels of intracellular sodium characteristic of this microorganism.

The unique properties of the A. luidlawii membrane were utilized by Steim et to show for the first time that biological membranes can undergo a gel-to-liquid-crystalline lipid phase transition similar to that previously reported for lamellar phospholipid-water systems. These workers demonstrated that when whole cells or isolated membranes are analyzed by differential scanning calorimetry (DSC), two relatively broad endothermic transitions are observed on the initial heating scan. The lowertemperature transition is fully reversible, varies markedly in position with changes in the length and degree of unsaturation of the membrane lipid fatty acyl chains, is broadened and eventually abolished by cholesterol incorporation, and exhibits a transition enthalpy characteristic of the mixed-acid synthetic phospholipids. Moreover, an endothermic transition having essentially identical properties is observed for the protein-free total membrane lipid extract dispersed in excess water or aqueous buffer, indicating that the presence of membrane proteins has little effect on the thennotropic phase behavior of most of the membrane lipids. The higher-temperature transition, in contrast, is irreversible, is independent on membrane lipid fatty acid composition or cholesterol content, and is absent in total membrane lipid extracts, indicating that it is due to an irreversible thermal denaturation of the membrane proteins. A comparison of the enthalpies of transition of the lipids in the membrane and in water dispersions indicates that at least 75% of the total membrane lipids participate in this transition. Evidence was also presented that the lipids must be predominantly in the fluid state to support normal growth. These results were later confirmed and extended by Reinert and Steim3 and by Melchior et al. ,4 who showed that the gel-to-liquid-crystalline

1. 1. The distribution of a number of fatty acids between the 1- and 2-positions of phosphatidyl glycerol from Mycoplasma laidlawii B has been investigated. The fatty acid composition of the phosphatidyl glycerol was systematically altered so that the positional distribution values determined here are accurate reflections of the relative positional affinities of the individual fatty acids examined. 2. 2. The positional distribution of any given fatty acid constituent was independent of the amount of that acid in the phosphatidyl glycerol, provided that the fatty acid in question was not present in sufficient quantity to saturate the preferred position. 3. 3. Within any single class of fatty acids, the positional specificity was markedly dependent on the chain length of the fatty acid. The smaller the total number of carbon atoms, the lesser was the affinity for the 1-position. 4. 4. Positional distribution was also dependent on the structural characteristics of the fatty acids. The following is the order of the fatty acid classes arranged according to a decreasing affinity for the 1-position: straight-chain saturated > branched-chain saturated > trans -monounsaturated > cis -monounsaturated > cyclopropane > cis -diunsaturated.

The fatty acid composition of the membrane polar lipids of Mycoplasma laidlawii B can be dramatically altered. These variations reslut in characteristic morphological chlanges, and in most cases the cells remain viable. This organism should provide a useful system for clarifying the role of fatty acyl chains in biological membranes.

Both membranes of Mycoplasma laidlawii and water dispersions of protein-free membrane lipids exhibit thermal phase transitions that can be detected by differential scanning calorimetry. The transition temperatures are lowered by increased unsaturation in the fatty acid residues, but in each case they are the same for membranes and lipids. The transitions resemble those observed for synthetic lipids in the lamellar phase in water, which arise from melting of the hydrocarbon chains within the phospholipid bilayers. Such melts are cooperative phenomena and would be greatly perturbed by apolar binding to protein. Thus the identity of membrane and lipid transition temperatures suggests that in the membranes, as in water, the lipids are in the bilayer conformation in which the hydrocarbon chains associate with each other rather than with proteins. Observations of morphological changes indicate that osmotic imbalance occurs when the membrane transition temperature exceeds the growth temperature, and that for transport processes to function properly the hydrocarbon chains must be in a liquid-like state.

1. 1. The fatty acid composition and cholesterol content of the membrane lipids of Acholeplasma laidlawii B were systematically altered and the rates at which glycerol and erythritol passively diffuse into intact cells and into liposomes prepared from the total membrane lipid were measured at a variety of temperature. 2. 2. The permeability of intact cells and derived liposomes is markedly dependent on the chemical structure and chain length of the fatty acids incorporated into the membrane lipids. Incorporation of branched-chain or unsaturated fatty acids, or fatty acids of reduced chain length, increases nonelectrolyte permeability to a similar extent in both cells and liposomes. 3. 3. The nonelectrolyte permeability of both the plasma and liposomal membrane is reduced by the incorporation of cholesterol. 4. 4. The mean activation energy values calculated for the permeation of glycerol and erythritol into intact cells are 18.2 and 21.3 kcal/mole, respectively. These values are the same, within experimental error, as those calculated for the liposomal system and suggest that glycerol and erythritol permeate both the biological and artificial membrane systems as single, fully dehydrate molecules. 5. 5. In contrast to permeation rates, which are dependent on both permeant structure and membrane lipid composition, activation energy values for the overall permeation process are dependent only on permeant structure and are not sgnificantly affected by variations in fatty acid composition or cholesterol content. 6. 6. The permeability of intact cells and derived liposomes is a function of the fluidity of the membrane lipids are measured by their reversible, thermotropic gel to liquid-crystalline phase transition temperatures. Cells or liposomes placed in isotonic permeant solutons undergo spontaneous lysis when the temperature is reduced to the point where most of the membrane lipids exist in the gel state.

We examined the permeabilization of lipid bilayers by the β-sheet, cyclic antimicrobial decapeptide gramicidin S (GS) in phospholipid bilayers formed either by mixtures of zwitterionic diphytanoylphosphatidylcholine and anionic diphytanoylphosphatidylglycerol or by single zwitterionic unsaturated phosphatidylcholines having various hydrocarbon chain lengths, with and without cholesterol. In the zwitterionic bilayers formed by the phosphatidylcholines, without or with cholesterol, the peptide concentrations and membrane potentials required to initiate membrane permeabilization vary little as function of bilayer thickness and cholesterol content. In all the systems tested, the GS-induced transient ion conductance events exhibit a broad range of conductances, which are little affected by the bilayer composition or thickness. In the zwitterionic phosphatidylcholine bilayers, the effect of GS does not depend on the polarity of the transmembrane potential; however, in bilayers formed from mixtures of phosphatidylcholines and anionic phospholipids, the polarity of the transmembrane potential becomes important, with the GS-induced conductance events being much more frequent when the GS-containing solution is positive relative to the GS-free solution. Overall, these results suggest that GS does not form discrete, well-defined, channel-like structures in phospholipid bilayers, but rather induces a wide variety of transient, differently sized defects which serve to compromise the bilayer barrier properties for small electrolytes.

The thermotropic phase behavior and organization bilayers composed of dimyristoyl phosphatidylethanolamine and some of its headgroup-modified analogues were investigated by differential scanning calorimetry and by fourier-transform infrared spectroscopy. As expected, chemical modification of the lipid headgroup resulted in changes in the gel/liquid-crystalline phase transition temperatures of the analogues that were positively correlated with the capacity for inter headgroup hydrogen-bonding to the phosphate moiety, and negatively correlated with the size of the headgroup substituent. However, such changes also drastically altered the nature of the lamellar crystalline phases formed by these compounds as well as the lateral packing interactions between the hydrocarbon chains. Moreover, although the chemical modifications of the polar headgroup did not alter the net charge of the lipid headgroup, they did nevertheless, markedly alter the susceptibility of these membranes to penetration by interfacially active molecules such as antimicrobial peptides. Our studies suggest that many of the effects of the chemical modification of the lipid headgroup can be rationalized in terms of their effects on the hydration of, and hydrogen-bonding interactions in the polar/polar interfacial regions of the lipid bilayer.

The thermotropic phase behavior and organization of model membranes derived from binary mixtures of tetramyristoylcardiolipin (TMCL) with dimyristoyl phosphatidylethanolamine (DMPE) and dimyristoyl phosphatidylglycerol (DMPG) were studied by differential scanning calorimetry (DSC) and fourier transform infrared (FTIR) spectroscopy. Cardiolipin-containing DMPE and DMPG model membranes all exhibit complex multi-component hydrocarbon chain-melting (Lβ/Lα) phase transitions upon heating and cooling. This suggests that TMCL is poorly miscible with both DMPE and DMPG and that the domains formed prior to the onset of the Lβ/Lα phase transitions are probably retained in the liquid-crystalline state. For all mixtures, the temperatures of the Lβ/Lα phase transitions are generally higher than those predicted by an ideal mixing of the components, and the enthalpies of these phase transitions seem to be better correlated with the component mol fraction of hydrocarbon chains than with the molar composition of the mixture per se. Finally, when cooled to low temperatures, cardiolipin-rich (i.e., > 30 mol % TMCL) membranes tend to form lamellar-crystalline phases which exhibit spectroscopic characteristics comparable to those exhibited by the lamellar crystalline phases of pure cardiolipin. Together, our experimental observations indicate that cardiolipin is poorly miscible with major lipid components of one of the biological membrane systems (Escherichia coli) in which it occurs naturally. The possibility that it may also be poorly miscible in the biologically relevant liquid-crystalline phase also has interesting structural and functional implications.Supported by the Canadian Institutes for Health Research, the Alberta Heritage Foundation for Medical Research and by the Consejo Nacional de Investigaciones Cientificas y Tecnicas (Argentina).

The peptides Aurein 1.2, Citropin 1.1, Maculatin 1.1 and Caerin 1.1 are members of four structurally related families of antimicrobial peptides produced in the skin secretions of Australian tree frogs. Although largely unstructured in aqueous solution, these peptides exhibit a high propensity for folding into amphipathic alpha helices when partitioned into lipid bilayers or dissolved in membrane mimetic media. Some of the distinguishing features of these families of antimicrobial peptides are that they usually form alpha helical structures with large hydrophobic surfaces (hydrophobic angle ∼200-240°), and the amino acid sequences of many of the larger members (i.e. those with sequence lengths >∼18 aa residues) usually contain significant amounts of helix-disrupting residues such as glycine and proline, the presence of which seems to be essential for the retention of antimicrobial activity. These helix-disrupting amino acid residues seem to be preferentially located in the C-terminal regions of the peptide where they tend to disrupt the break up the helical rod into two or more helical sections separated by disordered “flexible hinge” regions. The role of these “flexible hinges” has been the subject of considerable study and speculation. Our studies show that because of their large hydrophobic surfaces, they form helices with a high propensity for self association in aqueous media, and this property markedly diminishes the aqueous monomeric solubility of such peptides. Our results suggest the helix-disrupting amino acid residues may be essential for maintaining the aqueous solubility of these antimicrobial peptides