Impaired binding affinity of electronegative low-density lipoprotein (LDL) to the LDL receptor is related to nonesterified fatty acids and lysophosphatidylcholine content

Sonia Benítez, Virtudes Villegas, Cristina Bancells, Oscar Jorba, Francesc González-Sastre, Jordi Ordóñez-Llanos, José Luis Sánchez-Quesada

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Abstract

The binding characteristics of electropositive [LDL(+)] and electronegative LDL [LDL(-)] subfractions to the LDL receptor (LDLr) were studied. Saturation kinetic studies in cultured human fibroblasts demonstrated that LDL(-) from normolipemic (NL) and familial hypercholesterolemic (FH) subjects had lower binding affinity than their respective LDL(+) fractions (P < 0.05), as indicated by higher dissociation constant (KD) values. FH-LDL(+) also showed lower binding affinity (P < 0.05) than NL-LDL(+) (KD, sorted from lower to higher affinity: NL-LDL(-), 33.0 ± 24.4 nM; FH-LDL(-), 24.4 ± 7.1 nM; FH-LDL(+), 16.6 ± 7.0 nM; NL-LDL(+), 10.9 ± 5.7 nM). These results were confirmed by binding displacement studies. The impaired affinity binding of LDL(-) could be attributed to altered secondary and tertiary structure of apolipoprotein B, but circular dichroism (CD) and tryptophan fluorescence (TrpF) studies revealed no structural differences between LDL(+) and LDL(-). To ascertain the role of increased nonesterified fatty acids (NEFA) and lysophosphatidylcholine (LPC) content in LDL-(-), LDL(+) was enriched in NEFA or hydrolyzed with secretory phospholipase A2. Modification of LDL gradually decreased the affinity to LDLr in parallel to the increasing content of NEFA and/or LPC. Modified LDLs with a NEFA content similar to that of LDL(-) displayed similar affinity. ApoB structure studies of modified LDLs by CD and TrpF showed no difference compared to LDL(+) or LDL(-). Our results indicate that NEFA loading or phospholipase A2 lipolysis of LDL leads to changes that affect the affinity of LDL to LDLr with no major effect on apoB structure. Impaired affinity to the LDLr shown by LDL(-) is related to NEFA and/or LPC content rather than to structural differences in apolipoprotein B.
Original languageEnglish
Pages (from-to)15863-15872
JournalBiochemistry
Volume43
Issue number50
DOIs
Publication statusPublished - 21 Dec 2004

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