CRYSTALLOGRAPHIC STUDY OF THE STRUCTURE OF COLIPASE AND OF THE INTERACTION WITH PANCREATIC LIPASE

Colipase (M(r) 10 kDa) confers catalytic activity to pancreatic lipase under physiological conditions (high bile salt concentrations). Previ ously determined 3-Angstrom-resolution X-ray structures of lipase-coli pase complexes have shown that, in the absence of substrate, colipase binds to the noncatalytic C-terminal domain of pancreatic lipase (van Tilbeurgh H, Sarda L, Verger R, Cambillau C, 1992, Nature 359:159-162; van Tilbeurgh et al., 1993a, Nature 362:814-820). Upon lipid binding, conformational changes at the active site of pancreatic lipase bring a surface loop (the lid) in contact with colipase, creating a second b inding site for this cofactor. Covalent inhibition of the pancreatic l ipase by a phosphonate inhibitor yields better diffracting crystals of the lipase-colipase complex. From the 2.4-Angstrom-resolution structu re of this complex, we give an accurate description of the colipase. I t confirms the previous proposed disulfide connections (van Tilbeurgh H, Sarda L, Verger R, Cambillau C, 1992, Nature 359:159-162; van Tilbe urgh et al., 1993a, Nature 362:814-820) that were in disagreement with the biochemical assignment (Chaillan C, Kerfelec B, Foglizzo E, Chapu s C, 1992, Biochem Biophys Res Commun 184:206-211). Colipase lacks wel l-defined secondary structure elements. This small protein seems to be stabilized mainly by an extended network of five disulfide bridges th at runs throughout the flatly shaped molecule, reticulating its four f inger-like loops. The colipase surface can be divided into a rather hy drophilic part, interacting with lipase, and a more hydrophobic part, formed by the tips of the fingers. The interaction between colipase an d the C-terminal domain of lipase is stabilized by eight hydrogen bond s and about 80 van der Waals contacts. Upon opening of the lid, three more hydrogen bonds and about 28 van der Waals contacts are added, exp laining the higher apparent affinity in the presence of a lipid/water interface. The tips of the fingers are very mobile and constitute the lipid interaction surface. Two detergent molecules that interact with colipase were observed in the crystal, covering part of the hydrophobi c surface.