Life Sciences 70 (2001) 629–638 0024-3205/01/$ – see front matter © 2001 Elsevier Science Inc. All rights reserved. PII: S0024-3205(01)01439-4 Structure-function relationships of human apolipoprotein D An immunochemical analysis Laurence Terrisse a , Karine Marcoux a , Sonia Do Carmo a , Louise Brissette a , Ross Milne b , Eric Rassart a, * a Département des Sciences Biologiques, Université du Québec à Montréal, Montréal, Québec, Canada, H3C 3P8 b Lipoprotein and Atherosclerosis Research Group, University of Ottawa Heart Institute, 40 Ruskin St., Ottawa, Ontario, Canada, K1Y 4W79 Received 23 January 2001; accepted 26 June 2001 Abstract Apolipoprotein D (apoD), a 169 amino acid member of the lipocalin family, is thought to be a transporter of small, hydrophobic ligands. A panel of 10 anti-apoD monoclonal antibodies (mAbs) was prepared and characterized in order to define apoD structure-function relationships. An apoD epitope map was constructed based on reactivity of the mAbs with apoD fragments. Three mAbs react with epitopes between apoD residues 7–78, seven mAbs with epitopes between residues 128–169, one mAb recognizes an epitope that straddles residues 99–102 and one mAb is specific for an epitope composed of non-contiguous apoD residues. Several pairs of mAbs whose respective epitopes are widely sepa- rated in apoD primary structure can compete for binding to immobilized apoD. This would be consis- tent with the compact  -barrel tertiary structure that apoD is thought to adopt. None of the mAbs block the interaction of apoD with pregnenolone, a putative physiological ligand for apoD. © 2001 Elsevier Science Inc. All rights reserved. Keywords: Apolipoprotein D; Lipocalin; Epitope mapping; Transporter protein; Hydrophobic ligand; Mono- clonal antibody; Fusion protein Introduction Apolipoprotein D (apoD) is a 29 kDa glycoprotein that was originally identified as a com- ponent of high density plasma lipoproteins [1] and was subsequently shown to be identical to GCDFP-24, a progesterone-binding protein present in high concentration in mammary gross cystic fluid [2]. The cloning and sequencing of the human apoD cDNA revealed that the * Corresponding author. Dép. des Sciences Biologiques, Université du Québec à Montréal, C.P. 8888, succ. Centre-ville, Montréal, Québec, Canada, H3C 3P8. Tel.: 514 987-3000 Ext 3953#; fax: 514 987-4647. E-mail address: rassart.eric@uqam.ca (E. Rassart)