Four mycobacterial wall glycolipids were tested for their effects on phospholipidic liposome organization and passive permeability and on oxidative phosphorylation of isolated mitochondria. From fluorescence polarization of diphenylhexatriene performed on liposomes it was concluded that the two trehalose derivatives (dimycoloyltrehalose and polyphthienoyltrehalose) rigidified the fluid state of liposomes, the triglycosyl phenolphthiocerol slightly fluidized the gel state, while the peptidoglycolipid ("apolar" mycoside C) just shifted the phase transition temperature upward. Dimycoloyltrehalose was without effect on liposome passive permeability, as estimated from dicarboxyfluorescein leak rates, and polyphthienoyltrehalose and triglycosyl phenolphthiocerol slightly decreased leaks, while mycoside C dramatically increased leaks. Activity of these lipids on mitochondrial oxidative phosphorylation was examined. The two trehalose derivatives have been tested previously: both had the same type of inhibitory activity, dimycoloyltrehalose being the most active. Triglycosyl phenolphthiocerol was inactive. Mycoside C was very active, with effects resembling those of classical uncouplers: this suggested that its activity on mitochondria was related to its effect on permeability. All these membrane alterations were called nonspecific because it is likely that they result from nonspecific lipid-lipid interactions, and not from recognition between specific molecular structures. Such nonspecific interactions could be at the origin of some of the effects of mycobacteria glycolipids on cells of the immune system observed in the last few years.

Ionization of the acidic phospholipid phosphatidylglycerol has been studied by measuring the surface potential of monomolecular films of the lipid as a function of the aqueous subphase pH and the concentration of monovalent cations (Li, Na, Cs). It is shown that theexperimental data can be interpreted by means of the Gouy-Chapman theory in its simplest formulation, provided an adsorption of cations at the membrane surface is accounted for. This allows us to predict the ionization state of the lipid for given ionic conditions in the subphase. Above pH 4, for subphase ion concentration higher than 10 mM, or for ion concentrations above 0.1 mM at pH 5.6, phosphatidylglycerol is fully deprotonated. Within the limits of our theoretical approach, association constants of the cations to the lipid lie around 0.1-0.6 M-I.With the exception of phosphatidylcholine and phosphatidylethanolamine which are zwitterions over a large pH range (3 -8) [l], phospholipids in biomembranes are acidic substances bearing a net negative charge when they are ionized. Ionization or protonation of these acidic lipids as well as their interactions with cations can have considerable consequences on both membrane structure and functions.From a structural point of view, changes in the pH or the ionic strength of the aqueous phase have been shown to trigger phase transition of acidic phospholipids [2-111. The bivalent cations Mg2+ and Ca2+ can bring about phase separation within mixtures of zwitterionic and acidic phospholipids [12-191. Cations can also modify the dynamic properties of lipids, i.e. their lateral diffusion rate [20], as well as being able to promote morphological changes [21]. As an example, cardiolipin, which normally exists in a lamellar phase in the presence of sodium, adopts the hexagonal H,, phase in the presence of calcium [22, 231. Dipalmitoylglycerophosphoglycerol, which normally stays in a lamellar phase in the presence of sodium, has been shown to exist in an interdigitated phase in the presence of the organic cations choline and acetylcholine [24].From a functional point of view, phospholipids can modulate the activity of membrane enzymes by monitoring, at the membrane surface, a concentration of protons, cations and of any charged metabolites which is quite different from that existing in the bulk [25]. Charges at the membrane surface also contribute to the establishment of transmembrane potentials which are involved in energy-linked transmembrane transport processes [26] and which are postulated to monitor the orientation [27 -291 and the activity of certain membrane proteins [28 -341.For all these reasons, it is of great importance to have a full understanding of the ionization process of acidic phospholipids in membrane systems. This would allow a correct prediction of their ionization state and of the surface potential they generate, for given ionic conditions in the aqueous phase. As a first approach, ionization of phosphatidylglycerol was studied in monolayers by measuring changes in film surface pressure again...

In order to understand better the roles of repeating basic peptide motifs in modifying DNA structure, we have synthesized typical repeats found in the C-terminal domain of histone H I (KTPKKAKKP)2 and in the N-terminal domain of nucleolin (ATPAKKAA)2. By using circular dichroism in conjunction with Raman and Fouriertransform infrared spectroscopies, we demonstrate that the abilities of the two peptides to affect DNA conformation are dramatically different. Whilst the binding of the nucleolin repeat to DNA does not significantly alter its conformation, the binding of H I repeat induces a very marked DNA condensation, giving rise to a w( -)-type circular dichroic spectrum. The HI repeat thus adopts a more rigid p-turn-containing structure which probably binds to the DNA minor groove as assessed by competition with the drug Hoechst 33258. Unexpectedly, the DNA condensation induced by the H I repeat is enhanced by the nucleolin repeat which by itself does not promote any alteration in DNA conformation.Considerable effort has been devoted to characterizing the factors which regulate chromatin structure during the various stages of condensation and transcriptional activation. In particular, the manner in which chromatin DNA condensation is modulated in relation to nucleosomal organization remains an open question. From the outset, attention has been focused on one particular fragment of nucleosomal DNA, the socalled linker DNA, which ensures the tight junction between two adjacent compact core particles and is preferentially accessible to external factors such as nucleases. The folding of this DNA is under the control of the C-terminal domain of histone HI [l, 21. In consequence, histone H1 and all the nonhistone proteins which can interact or compete with HI are likely to play an important role in regulating local access to the linker DNA.In a previous report 131, we have shown that nucleolin, a major nucleolar protein, can decondense chromatin by binding to histone H1. Indeed, the bipolar composition of the nucleolin N-terminal domain is analogous to that of the highmobility-group (HMG) proteins. The presence of long acidic stretches probably explains its ability to bind to histone H1 and this binding leads to an unfolding of chromatin linker DNA. We have concluded that nucleolin most likely displaces HI, or at least the C-terminal domain of HI, from its interaction with linker DNA. However, these acidic stretches are also interspersed with a basic and repeated octapeptide motif [3, 41 XTPXKKXX (where X designates a non-polar residue) which could be responsible for the capacity of nucleolin to bind to chromatin 151. As we have already pointed out [3] disposition of lysine residues within a short repeated sequence is reminiscent of that present in the HI C-terminal domain where one finds various arrangements of repeating oligopeptides containing mostly lysine, alanine, proline and serine or threonine 16-81, (for a review see, [9]). The compilation and comparison of the sequences of many histone H1 proteins [lo, 111 ...