title: Molecular modeling of the dimeric structure of human lipoprotein lipase and functional studies of the carboxyl-terminal domain.
abstract: Lipoprotein lipase (LPL) plays a key role in lipid metabolism. Molecular modeling of dimeric LPL was carried out using insight ii based upon the crystal structures of human, porcine, and horse pancreatic lipase. The dimeric model reveals a saddle-shaped structure and the key heparin-binding residues in the amino-terminal domain located on the top of this saddle. The models of two dimeric conformations - a closed, inactive form and an open, active form - differ with respect to how surface-loop positions affect substrate access to the catalytic site. In the closed form, the surface loop covers the catalytic site, which becomes inaccessible to solvent. Large conformational changes in the open form, especially in the loop and carboxyl-terminal domain, allow substrate access to the active site. To dissect the structure-function relationships of the LPL carboxyl-terminal domain, several residues predicted by the model structure to be essential for the functions of heparin binding and substrate recognition were mutagenized. Arg405 plays an important role in heparin binding in the active dimer. Lys413/Lys414 or Lys414 regulates heparin affinity in both monomeric and dimeric forms. To evaluate the prediction that LPL forms a homodimer in a 'head-to-tail' orientation, two inactive LPL mutants - a catalytic site mutant (S132T) and a substrate-recognition mutant (W390A/W393A/W394A) - were cotransfected into COS7 cells. Lipase activity could be recovered only when heterodimerization occurred in a head-to-tail orientation. After cotransfection, 50% of the wild-type lipase activity was recovered, indicating that lipase activity is determined by the interaction between the catalytic site on one subunit and the substrate-recognition site on the other.
authors.affiliation: Division of Genetic Diagnosis, Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
authors.forename: Yoko
authors.initials: Y
authors.lastname: Kobayashi
authors.forename: Toshiaki
authors.initials: T
authors.lastname: Nakajima
authors.forename: Ituro
authors.initials: I
authors.lastname: Inoue
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D002522:Chlorocebus aethiops; D019281:Dimerization; D006493:Heparin / Q000378:metabolism; D006801:Humans; D008071:Lipoprotein Lipase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D011487:Protein Conformation; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary
chemical_list: D006493:Heparin; D008071:Lipoprotein Lipase