Acylated and unacylated ghrelin binding to membranes and to ghrelin receptor: towards a better understanding of the underlying mechanisms.
The O-octanoylation of human ghrelin is a natural post-translational modification that enhances its binding to model membranes and could potentially play a central role in ghrelin biological activities. Here, we aimed to clarify the mechanisms that drive ghrelin to the membrane and hence to its receptor that mediates most of its endocrinological effects. As the acylation enhances ghrelin lipophilicity and that ghrelin contains many basic residues, we examined the electrostatic attraction and/or hydrophobic interactions with membranes. Using various liposomes and buffer conditions in binding, zeta potential and isothermal titration calorimetry studies, we found that whereas acylated and unacylated ghrelin were both electrostatically attracted towards the membrane, only acylated ghrelin penetrated into the headgroup and the lipid backbone regions of negatively charged membranes. The O-acylation induced a 120-fold increase in ghrelin local concentration in the membrane. However, acylated ghrelin did not deeply penetrate the membrane nor did it perturb its organisation. Conformational studies by circular dichroism and attenuated total reflection Fourier transformed infrared as well as in silico modelling revealed that both forms of ghrelin mainly adopted the same structure in aqueous, micellar and bilayer environments even though acylated ghrelin structure is slightly more α-helical in a lipid bilayer environment. Altogether our results suggest that membrane acts as a "catalyst" in acylated ghrelin binding to the ghrelin receptor and hence could explain why acylated and unacylated ghrelin are both full agonists of this receptor but in the nanomolar and micromolar range, respectively.
Staes
Edith
E
Université catholique de Louvain, Unité de Pharmacie Galénique, 1200 Brussels, Belgium. edith.staes@gmail.com
Absil
Pierre-Antoine
PA
Lins
Laurence
L
Brasseur
Robert
R
Deleu
Magali
M
Lecouturier
Nathalie
N
Fievez
Virginie
V
Rieux
Anne des
Ad
Mingeot-Leclercq
Marie-Paule
MP
Raussens
Vincent
V
Préat
Véronique
V
D000215:Acylation; D005260:Female; D054439:Ghrelin / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008297:Male; D008560:Membrane Fluidity; D011487:Protein Conformation; D021381:Protein Transport; D054440:Receptors, Ghrelin / Q000737:chemistry / Q000378:metabolism*; D055672:Static Electricity
D054439:Ghrelin; D054440:Receptors, Ghrelin