Missense mutation (Gly----Glu188) of human lipoprotein lipase imparting functional deficiency

Cloning and sequencing of lipoprotein lipase (LPL) cDNA prepared from the adipose tissue of a patient with classical LPL deficiency revealed a G to A transition at nucleotide 818 in all sequenced clones, leading to the substitution of glutamic acid for glycine at residue 188 of the mature protein. Hybridization of genomic DNA with allele-specific oligonucleotides confirmed that the patient was homozygous for this mutation and revealed that carrier status for this mutation among relatives of the patient was significantly associated with hypertriglyceridemia. Assay of the patient's plasma for immunoreactive enzyme and activity demonstrated the presence of a circulating inactive enzyme protein, the concentration of which was further increased by injection of heparin. The mutant sequence was produced by oligonucleotide-directed mutagenesis, and both normal and mutant sequences were cloned into the expression vector pSVL and transfected into COS-1 cells. The normal sequence led to the in vitro expression of an enzyme that bound to heparin-Sepharose and had a specific catalytic activity similar to that of normal postheparin plasma enzyme. By contrast, the mutant enzyme expressed in vitro was catalytically inactive and displayed a lower affinity for heparin than the normal enzyme. We conclude that this single amino acid substitution leads to the in vivo expression of an inactive enzyme accounting for the manifestations of LPL deficiency noted in the patient.