Deletion of the helical motif in the intestinal fatty acid-binding proteinreduces its interactions with membrane monolayers: Brewster angle microscopy, IR reflection-absorption spectroscopy, and surface pressure studies

Intestinal fatty acid binding protein (IFABP) appears to interact directly with membranes during fatty acid transfer [Hsu, K. T., and Storch, J. (1996 ) J. Biol. Chem. 271, 13317-13323]. The largely a-helical "portal" domain o f IFABP was critical for these protein-membrane interactions. In the presen t studies, the binding of IFABP and a helixless variant of IFABP (IFABP-HL) to acidic monolayers of 1,2-dimyristoylphosphatidic acid (DMPA) has been m onitored by surface pressure measurements, Brewster angle microscopy (BAM), and infrared reflection-absorption spectroscopy (LRRAS). Protein adsorptio n to DMPA exhibited a two phase kinetic process consisting of an initial sl ow phase, arising from protein binding to the monolayer and/or direct inter facial adsorption, and a more rapid phase that parallels formation of lipid -containing domains. IFABP exhibited more rapid changes in both phases than IFABP-HL. The second phase was absent when IFABP interacted with zwitterio nic monolayers of 1,2-dipalmitoylphosphatidylcholine, revealing the importa nt role of electrostatics at this stage. BAM images of DMPA monolayers with either protein revealed the formation of domains leading eventually to rig id films. Domains of DMPA/IFABP-HL formed more slowly and were less rigid t han with the wild-type protein. Overall, the IRRAS studies revealed a prote in-induced conformational ordering of the lipid acyl chains with a substant ially stronger ordering effect induced by IFABP. The physical measurements thus suggested differing degrees of direct interaction between the proteins and DMPA monolayers with the IFABP/ DMPA interaction being somewhat strong er. These data provide a molecular structure rationale for previous kinetic measurements indicating that the helical domain is essential for a collisi on-based mechanism of fatty acid transfer to phospholipid membranes [Corsic o, B., Cistola, D. P., Frieden, C. and Storch, J. (1998) Proc. Natl. Acad. Sci. U.S.A. 95, 12174-12178].