Membrane penetration depth of Intestinal Fatty Acid Binding Protein (IFABP)

EDUARDO DE GERÓNIMO; JUDITH STORCH; BETINA CÓRSICO

Bilbao, España.

Congreso; 51st International Conference on the Bioscience of Lipids; 2010

Intestinal FABP (IFABP) belongs to a family of small cytosolic proteins involved in lipid transport and metabolism that share a common structure. IFABP structure consists of ten antiparallel β-strands that form a β-barrel, capped by two short α-helices. The helical domain is postulated to be part of a small portal at the top of the barrel, wherein a hydrophobic ligand enters and exits the cavity of the barrel. Fatty acid transfer from IFABP to phospholipid membranes is proposed to occur during protein-membrane collisional interactions. Employing a helix-less variant of IFABP, we have shown that the α-helical region of IFABP is involved in membrane interactions, and appears to play a primary role in the collisional mechanism of fatty acid transfer from IFABP to membranes. The objective of this study was to determine the membrane penetration depth of the IFABP portal region by fluorescence quenching techniques and analyze how it is affected by the charge of the membrane surface and by the ionic strength. We have engineered a series of IFABP single-tryptophan mutants in the portal region. The Trp fluorescence of these mutants is quenched by brominated phosphatidylcholine localized in large unilamellar vesicles, with bromines at different positions along the fatty acid acyl chain. The results clearly show that the IFABP portal region is the membrane binding domain and is located within the lipid bilayer. Moreover, increasing the negative charge of the membrane could affect the depth and angle of insertion depth.