Conformational rearrangement of beta-lactoglobulin upon interaction with an anionic membrane

Interactions between P-lactoglobulin (beta -1g) and dimyristoylphosphatidyl glycerol (DMPG) bilayers were studied using one and two-dimensional infrare d spectroscopy above (pD 7.4) and below (pD 4.4) the protein's (beta -lg's) isoelectric point (pI= 5.2). The aim of the study was threefold: (1) gain a better understanding of beta -lg-phospholipid interaction, (2) provide in formation relative to the structure of beta -lg as it interacts with membra nes, (3) determine whether the conformational modifications of the protein in the presence of lipids are strictly caused by thermal effects or whether they are modulated by the chain-melting phase transition. At pD 7.4 the li pid thermotropism, the acyl-chain order, and the membrane interfacial regio n were essentially unaffected by the presence of beta -lg, whereas the prot ein amide I region showed dramatic alterations. The results suggested the p redominance of beta -sheets and (x-helix elements, with a lost of structura l integrity. At pD 4.4. P-Ig induced a similar to 2 degreesC downshift of t he transition temperature, whereas the conformational order of the lipid ch ain decreased in the gel phase and increased in the liquid-crystalline phas e. The hydration state of the DMPG C = O groups increased in the liquid-cry stalline phase. The conformation of beta -lg at pD 4.4 in the presence of D MPG showed similarities with that observed at pD 7.4, but an increase in th e oc-helix content and a reduced thermal stability were noticed. In contras t to the protein alone, beta -lg aggregates in the presence of DMPG at pD 4 .4 above 50 degreesC. At both pD values, the charged surface of the membran e seemed to be the main factor for inducing protein conformational changes by altering the intramolecular interactions that stabilize the native struc ture. However, protein incorporation within the membrane seemed to be invol ved at pD 4.4. The two-dimensional analysis performed with spectra recorded upon heating showed that spectral intensity changes at pD 4.4 and 7.4 occu rred at the same frequencies in the amide I ' region. The heat-induced stru ctural changes of P-Ig were not correlated with the conformational modifica tions of the phospholipids along the phase transition, indicating that the thermal behavior of the protein was not modulated by the lipid chain meltin g, but rather represented the heat-induced protein rearrangement in the pre sence of DMPG. (C) 2001 Elsevier Science BN. All rights reserved.