Membrane-promoted unfolding of acetylcholinesterase: A possible mechanism for insertion into the lipid bilayer

Abstract

Acetylcholinesterase from Torpedo californica partially unfolds to a state with the physicochemical characteristics of a ''molten globule'' upon mild thermal denaturation or upon chemical modification of a single nonconserved buried cysteine residue, Cys 231 . The protein in this state binds tightly to liposomes. It is here shown that the rate of unfolding is greatly enhanced in the presence of unilamellar vesicles of dimyristoylphosphatidylcholine, with concomitant incorporation of the protein into the lipid bilayer. Arrhenius plots reveal that in the presence of the liposomes the energy barrier for transition from the native to the molten globule state is lowered from 145 to 47 kcal/mol. Chemical modification of Cys 231 by mercuric chloride produces initially a quasinative state of Torpedo acetylcholinesterase which, at room temperature, undergoes spontaneous transition to a molten globule state with a half-life of 1-2 hr. This permitted temporal resolution of interaction of the quasi-native state with the membrane from the transition of the membrane-bound protein to the molten globule state. The data presented here suggest that either the native enzyme, or a quasi-native state with which it is in equilibrium, interacts with the liposome, which then promotes a fast transition to the membrane-bound molten globule state by lowering the energy barrier for the transition. These findings raise the possibility that the membrane itself, by lowering the energy barrier for transition to a partially unfolded state, may play an active posttranslational role in insertion and translocation of proteins in situ.

The role of lipid–protein interactions in amyloid-type protein fibril formation

“…An increasing number of studies provides support to the idea that lipid bilayers can lower the activation energy barrier for protein unfolding. Partial unfolding in a membrane environment has been reported for cytochrome c (Muga et al, 1991;Pinheiro et al, 1997), phospholipase A 2 (Tatulian et al, 1997), bacterial toxins (Butko et al, 1997;Muga et al, 1993), acetylcholinesterase (Shin et al, 1997), pheromone-binding protein (Wojtasek and Leal, 1999), and recombinant human prion protein (Morillas et al, 1999). Conformational transitions are usually interpreted as originating from the lowered interfacial pH, being a consequence of proton accumulation in the vicinity of negatively charged membrane surface (Träuble, 1977).…”

Membrane Environment Alters the Conformational Structure of the Recombinant Human Prion Protein

“…However, the effect described in this study is not unprecedented. Membrane surface-induced destabilization and/or partial unfolding have been recently reported for few other proteins including cytochrome c (32,33), acetylcholinesterase (34), phospholipase A 2 (35), and some bacterial toxins (27,36).…”

“…At present, no data are available about the biophysical properties of the membrane-bound prion protein. Information in this regard is of potentially critical importance since studies with other membrane-interacting proteins indicate that the specific environment at the membrane surface may have a profound influence on a protein, affecting its conformational structure, folding, and stability (27,(32)(33)(34)(35)(36).…”

Lipid-assisted Protein Folding

“…The liposome-induced conformational transition of colicin E1 occurs concomitantly with insertion into membranes (30,31). The rate of unfolding of acetylcholine esterase from Torpedo californica was greatly enhanced in the presence of PC vesicles with concomitant insertion of the protein into the lipid bilayer (32). For ␤-barrel membrane proteins, folding and membrane insertion are coupled processes that involve kinetically distinguishable steps (10,33).…”