Mechanism of LDL binding and release probed by structure-based mutagenesis of the LDL receptor.
The LDL receptor (LDL-R) mediates cholesterol metabolism in humans by binding and internalizing cholesterol transported by LDL. Several different molecular mechanisms have been proposed for the binding of LDL to LDL-R at neutral plasma pH and for its release at acidic endosomal pH. The crystal structure of LDL-R at acidic pH shows that the receptor folds back on itself in a closed form, obscuring parts of the ligand binding domain with the epidermal growth factor (EGF)-precursor homology domain. We have used a structure-based site-directed mutagenesis approach to examine 12 residues in the extracellular domain of LDL-R for their effect on LDL binding and release. Our studies show that the interface between the ligand binding domain and the EGF-precursor homology domain seen at acidic pH buries residues mediating both LDL binding and release. Our results are consistent with an alternative model of LDL-R whereby multiple modules of the extracellular domain interact with LDL at neutral pH, concurrently positioning key residues so that at acidic pH the LDL-R:LDL interactions become unfavorable, triggering release. After LDL release, the closed form of LDL-R may target its return to the cell surface.
Huang
Sha
S
Life Sciences Institute, University of Michigan, Ann Arbor, Michigan, USA.
Henry
Lisa
L
Ho
Yiu Kee
YK
Pownall
Henry J
HJ
Rudenko
Gabby
G
D018360:Crystallography, X-Ray; D004815:Epidermal Growth Factor / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008077:Lipoproteins, LDL / Q000378:metabolism*; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D011973:Receptors, LDL / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship
D008077:Lipoproteins, LDL; D011973:Receptors, LDL; D004815:Epidermal Growth Factor
Science. 2002 Dec 20;298(5602):2353-8
12459547
Science. 1988 Apr 29;240(4852):622-30
3283935
Semin Vasc Med. 2004 Feb;4(1):5-11
15199428
Science. 2006 Mar 24;311(5768):1721-3
16556829
J Clin Invest. 1998 Mar 1;101(5):1084-93
9486979
Science. 2001 May 18;292(5520):1310-2
11360986
Proc Natl Acad Sci U S A. 2008 Sep 2;105(35):13045-50
18753623
Biochemistry. 2008 Nov 4;47(44):11647-52
18847225
J Cell Sci. 2005 Mar 15;118(Pt 6):1309-20
15741231
Nature. 1987 Apr 23-29;326(6115):760-5
3494949
J Biol Chem. 2008 Aug 15;283(33):22670-9
18574243
Nature. 1997 Aug 14;388(6643):691-3
9262405
Nat Struct Biol. 2001 Jun;8(6):499-504
11373616
Biochemistry. 2005 Apr 5;44(13):5075-85
15794645
Biochemistry. 2001 Dec 18;40(50):15127-34
11735395
Science. 1985 Oct 4;230(4721):37-43
2994225
Methods Enzymol. 1983;98:241-60
6321901
Curr Biol. 1998 Dec 3;8(24):1335-8
9843688
J Cell Sci. 2002 Feb 1;115(Pt 3):599-607
11861766
Ann Hum Genet. 2008 Jul;72(Pt 4):485-98
18325082
J Biol Chem. 1989 Oct 5;264(28):16584-90
2777800
Mol Cell. 2006 Apr 21;22(2):277-83
16630895
Proc Natl Acad Sci U S A. 1974 Mar;71(3):788-92
4362634
J Biol Chem. 1989 Dec 25;264(36):21682-8
2600087
Biochem J. 1991 Aug 1;277 ( Pt 3):677-82
1872803
Mol Cell. 2004 Oct 22;16(2):281-92
15494314
Curr Opin Struct Biol. 2003 Dec;13(6):683-9
14675545
J Biol Chem. 1988 Sep 15;263(26):13282-90
3417658
Cell. 1984 Nov;39(1):27-38
6091915
J Mol Biol. 2006 Sep 29;362(4):700-16
16938309
J Biol Chem. 2008 Sep 26;283(39):26528-37
18677035
J Lipid Res. 2006 Jul;47(7):1399-405
16601300
Hum Mutat. 2002 Aug;20(2):81-7
12124988
J Biol Chem. 1989 Jun 5;264(16):9533-8
2722848