Apolipoprotein H (ApoH) is a plasma glycoprotein with its in vivo physiological and pathogenic roles being closely related to its interaction with negatively charged membranes. In this paper, the interaction of ApoH with phospholipid vesicles was characterized by (i) detecting the wavelength shift of the fluorescence spectrum of ApoH and (ii) measuring the fluorescence quenching extent of ApoH by the membrane resident quencher 1-palmitoyl-2-stearoyl-(5-doxyl)-sn-glycero-3-phosphocholine (DPC). The observed blue shift upon addition of DMPG vesicles indicated that the tryptophan residues of ApoH moved from a polar to a nonpolar environment. The insertion ability of ApoH into PG-containing v esicles did not depend on the PG content in a stoichiometric way as did the blue shift, indicating that the negatively charged DMPG does not serve as a specific binding site but rather provides a suitable microenvironment for ApoH interaction. The finding that the detachment effect of cations on the blue shift is remarkably different from that on the quenching extent suggests that ApoH is capable of existing in two different conformations when membrane-bound.

Trichosanthin (TCS) is the active component extracted from Tianhuafen, a traditional herbal medicine that has been used for abortion in China for centuries. It belongs to the type-I ribosomeinactivating protein (RIP) family and can inactivate the eukaryotic ribosome through its RNA N-glycosidase activity. Recentstudies have shown TCS to be multifunctional, its pharmacologicalproperties including immunomodulatory, anti-tumourand anti-HIV activities. The membrane-insertion property ofTCS is thought to be essential for its physiological effect, for itmust get across the membrane before it can enter the cytoplasmand exert its RIP function. In this paper, the membrane-insertionmechanism of TCS was studied. The monolayer experimentrevealed that TCS's membrane-insertion ability was dependenton low pH. Fluorescence spectroscopy using 1-anilinonaph-thalene-8-sulphonic acid as a probe showed that low pH mayinduce the conformational change of TCS that leads to thehydrophobic-site exposure, and the CD result showed that thisconformational change did not alter its secondary structure.Such conformational change leads to an intermediate state,called the molten globular state' by previous investigators. ThepH-dependent membrane insertion and conformational changewere related by the fact that the optimal membrane-surface pHneeded was the same for the two events. From these and otherresults, a membrane-insertion model was proposed.

Apolipoprotein H (ApoH), also known as β2-glycoprotein I, is a plasma glycoprotein with its in vivo physiological and pathogenic roles being closely related to its interaction with negatively charged membranes. Although the threedimensional crystal structure of ApoH has been recently solved, direct evidence about the spatial state of ApoH on the membrane is still lacking. In this work, the interactions of ApoH with the lipid layer are studied by a combination of lipid monolayer approach and surface concentration determination. The spatial state of the orientation of ApoH on the lipid layer is investigated by analyzing the process of membrane-attached ApoH molecules being extruded out from the phospholipid monolayer by compression. The results show that on neutral lipid layer ApoH has an upright orientation, which is not sensitive to the phase state of the lipid layer. However, on acidic lipid layer, ApoH may have two forms of orientation. One is an upright orientation in the liquid phase region, and the other is flat orientation on the condensed domain region. The variation of the spatial state of ApoH on the lipid layer may relate to a variety of its physiological functions.

The interaction of apolipoprotein H (Apo H) with lipid membrane has been considered to be a basic mechanism for the biological function of the protein. Previous reports have demonstratedthat Apo H can interact only with membranes containinganionic phospholipids. Here we study the membrane-inducedconformational change of Apo H by CD spectroscopy with twodifferent model systems: anionic-phospholipid-containing liposomes[such as 1,2-dimyristoyl-sn-glycero-3-phosphoglycerol(DMPG) and cardiolipin], and the water}methanol mixtures atmoderately low pH, which mimic the micro-physicochemicalenvironment near the membrane surface. It is found that Apo Hundergoes a remarkable conformational change on interactionwith liposomes containing anionic phospholipid. To interact withliposomes containing DMPG, there is a 6.8%increase in a-helixin the secondary structures; in liposomes containing cardiolipin,however, there is a 12.6% increase in a-helix and a 9% decreasein b-sheet. The similar conformation change in Apo H can beinduced by treatment with an appropriate mixture of water}methanol. The results indicate that the association of Apo Hwith membrane is correlated with a certain conformationalchange in the secondary structure of the protein.