STRUCTURAL STUDIES OF D98D, A TRUNCATED BUT FUNCTIONAL FORM OF INTESTINAL FATTY ACID BINDING PROTEIN (IFABP)

CURTO L.M.; DELFINO J.M.

San Carlos de Bariloche, Río Negro. Argentina

Congreso; Bariloche Protein Symposium, XXXIX Reunion Anual de la Sociedad Argentina de Investigacion Bioquimica y Biologia Molecular (SAIB), XXXII Reunion Anual de la Sociedad Argentina de Biofisica (SAB).; 2003

IFABP is a 15kDa intracellular lipid binding protein exhibiting a b-clam structure built of 2 perpendicular 5-stranded b-sheets and an intervening helix-turn-helix motif located in between strands A and B. D98D is the main product obtained after limited proteolysis of IFABP with clostripain. This fragment (29-126) lacks b-strand A, most of the helical domain, and also the last 5 amino acids belonging to the C-terminal b-strand. Despite this, it remains stable and soluble. D98D is purified in its apo form and free from any other proteolytic fragment. CD spectroscopy shows significant b-sheet content while near UV absorption and intrinsic fluorescence emission spectra reveal an identical tryptophan environment: <l>em 341.3nm for both IFABP and D98D. These results are consistent with the conservation in D98D of all the hydrophobic amino acid residues (47,62,64,68,82,84 and 89) implicated in the nucleation event leading to the folded state. The fact that the Stokes radius of D98D is ~30% larger than that expected from its molecular weight (11kDa) and that this fragment unfolds less cooperatively and at a lower urea concentration (3.1 M) than IFABP, lends support to the view that D98D adopts a folded non-compact state. D98D is the smallest structure of this kind described so far preserving structure and binding activity.