pH-dependent Way Research Articles

Design, synthesis and structure of an amphipathic peptide with pH-inducible haemolytic activity.

A synthetic, 26-residue peptide having a strong helix forming potential in the protonated state was designed to interact with lipid bilayers in a pH-dependent way. On the basis of this concept a cluster of four glutamic acid residues was inserted in the central region of the amphipathic peptide to promote helix destabilization by mutual charge repulsion at neutral pH. Protonation of these residues might then bring about both a pH-mediated change in hydrophobicity and conformation forming a membrane-active amphiphilic helix. The sequence GLGTLLTLLEFLLEELLEFLKRKRQQamide produced by the design strategy induced pH-triggered lysis of human erythrocytes. A molecular model correlating the lytic activity to the formation of transmembrane pores which were detected by electron microscopy in erythrocyte membranes is discussed. Circular dichroism studies indicated a self-association of the monomeric random coil form with increasing peptide concentration leading to the apparent induction of strong alpha-helix formation (approximately 100% helicity) in the fully aggregated state. However, no pH-dependent helix-random coil transition was observed, implying that interhelical hydrophobic and ionic interactions not only govern the self-association but also decisively influence the conformational stability of the peptide.

The anti-invasive flavonoid (+)-catechin binds to laminin and abrogates the effect of laminin on cell morphology and adhesion

To study the effect of the flavonoid (+)-catechin on cell-matrix interactions two cell types with a different morphology on and adhesion to laminin were used. MO 4 virally transformed fetal mouse cells adhere and spread when cultured on top of laminin-coated coverslips or on human amnion basement membrane. M5076 mouse reticulum cell sarcoma cells poorly adhere to these substrates and remain round. Both cell types are invasive in confronting cultures with embryonic chick heart fragments. (+)-Catechin binds to laminin in a pH-dependent way. Pretreatment of laminin-coated coverslips or amnion basement membrane with 0.5 m M (+)-catechin abrogates the effect of laminin on cell morphology and adhesion. MO 4 cells do not adhere to the pretreated substrates and remain round, while M5076 cells now adhere and spread. (+)-Catechin inhibits the invasion of MO 4 cells but not of M5076 cells into embryonic chick heart in vitro. We speculate that the anti-invasive activity of the flavonoid to MO 4 cells is the result of its interference with MO 4 cell adhesion to laminin. Invasion of M5076 cells does not imply adhesion to and spreading on laminin.

Effects of pH and ascorbic acid on 3H-spiperone and 3H-dihydroalprenolol binding in rat forebrain homogenates.

The binding of the neuroleptic agent 3H-spiperone and the beta-blocker 3H-dihydroalprenolol (3H-DHA) was investigated in rat forebrain homogenates. The pH of the buffer medium was found to specifically affect binding to certain sites. Omission of ascorbic acid from the binding assay influenced binding in a pH-dependent way. Preincubation with a relatively low concentration of ascorbic acid markedly reduced the binding of both ligands except the nonspecific binding of 3H-spiperone. Lipid peroxidation as revealed by malonyldialdehyde formation, was increased by low and decreased by high ascorbic acid concentrations. The possible relationship between changes in number of binding sites and liposolubility is discussed.

The role of divalent metal ions in the mechanism of action of the hormone angiotensin-II

The pressor and myotropic activities in vivo of the linear octapeptide hormone Angiotensin-II (A-II) (Asp-Arg-Val-Tyr-Ile-His-Pro-Phe) are enchanced by divalent metal ions. In model membrane systems consisting of phospholipid vesicles, A-II behaves as ionophore in translocating Mn(II) ions across the bilayer [1] with a rate constant which is pH dependent with an apparent pK of 7.7. Since A-II is itself interacting with the bilayer in pH dependent way [2], the metal-A-II coordination modes are expected to play a central role in the biological effect of the hormone. In this report nuclear and electron relaxation features of the A-IIMn(II) complexes in aqueous solution are measured with the aim of delineating the metal binding sites. The ESR intensity loss and the appearance of ‘slow motion’ features in the X-band ESR spectrum of Mn(II) were taken to demonstrate a main binding interaction with one (or two) carboxyl groups, since the ESR parameters were pH independent over the whole range investigated (pH = 4–8). The paramagnetic contribution to nuclear relaxation rates. T −1 ip, were measured for aromatic protons of A-II: (a typical spectrum is shown in Fig. 1) and, since T −1 2p < T −1 1p for all the protons, i) fast exchange conditions were assumed and ii) dipolar only contributions to T −1 1p were considered. The T −1 1p and T −1 2p values were therefore taken to demonstrate that the imidazole moiety of His 6 is a secondary metal binding site. The pH dependence of T −1 1p, reported in Table I, clearly showed that the binding to His 6 is relevant in a narrow pH range only, which could be explained in t001 I:pH dependence of T −1 1p for Phe and His H 2 protons of A-II. [A-II] = 0.01 M; [Mn ++] = 0.01 m M; T = 295 K. pH T 1p −1 (sec −1 Phe His H 2 4.5 0.12 0.14 5.5 0.18 0.62 6.0 0.22 1.83 6.5 0.24 6.22 7.0 0.22 4.90 7.5 0.19 2.05 8.0 0.14 1.62 ▪ terms of intramolecular structuring within the peptide molecule opposing metal coordination driven by deprotonation of the imidazole ring.