Low-density lipoprotein receptor is a calcium/magnesium sensor - role of LR4 and LR5 ion interaction kinetics in low-density lipoprotein release in the endosome.
The low-density lipoprotein receptor (LDLR) captures circulating lipoproteins and delivers them in the endosome for degradation. Its function is essential for cholesterol homeostasis, and mutations in the LDLR are the major cause of familiar hypercholesterolemia. The release of LDL is usually attributed to endosome acidification. As the pH drops, the affinity of the LDLR/LDL complex is reduced, whereas the strength of a self-complex formed between two domains of the receptor (i.e. the LDL binding domain and the β-propeller domain) increases. However, an alternative model states that, as a consequence of a drop in both pH and Ca(2+) concentration, the LDLR binding domain is destabilized in the endosome, which weakens the LDLR/LDL complex, thus liberating the LDL particles. In the present study, we test a key underlying assumption of the second model, namely that the lipoprotein binding repeats of the receptor (specifically repeats 4 and 5, LR4 and LR5) rapidly sense endosomal changes in Ca(2+) concentration. Our kinetic and thermodynamic analysis of Ca(2+) and Mg(2+) binding to LR4 and LR5, as well as to the tandem of the two (LR4-5), shows that both repeats spontaneously release Ca(2+) in a time scale much shorter than endosomal delivery of LDL, thus acting as Ca(2+) sensors that become unfolded under endosomal conditions. Our analysis additionally explains the lower Ca(2+) affinity of repeat LR4, compared to LR5, as arising from a very slow Ca(2+) binding reaction in the former, most likely related to the lower conformational stability of apolipoprotein LR4, compared to apolipoprotein LR5, as determined from thermal unfolding experiments and molecular dynamics simulations.
Biocomputation and Complex Systems Physics Institute (BIFI) - Joint Unit BIFI-IQFR (CSIC), Universidad de Zaragoza, Spain; Departamento de Bioquímica y Biología Molecular y Celular, Universidad de Zaragoza, Spain.
Juan
J
Martínez-Oliván
Zuriñe
Z
Rozado-Aguirre
Xabier
X
Arias-Moreno
Vladimir E
VE
Angarica
Adrián
A
Velázquez-Campoy
Javier
J
Sancho
D002118:Calcium / Q000378:metabolism*; D011992:Endosomes / Q000378:metabolism; D005453:Fluorescence; D007700:Kinetics; D008077:Lipoproteins, LDL / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism*; D008962:Models, Theoretical; D056004:Molecular Dynamics Simulation; D011485:Protein Binding; D011973:Receptors, LDL / Q000235:genetics / Q000378:metabolism*
D008077:Lipoproteins, LDL; D011973:Receptors, LDL; D008274:Magnesium; D002118:Calcium
Ca2+ binding; ITC; fluorescence; kinetics; low-density lipoprotein receptor