Characterization of the calcium site in two complement-like domains from the low-density lipoprotein receptor-related protein (LRP) and comparison with a repeat from the low-density lipoprotein receptor.
Calcium is required for the binding and endocytosis of protein ligands by the low-density lipoprotein receptor-related protein (LRP) and other members of the low-density lipoprotein (LDL) receptor family. Calcium binding sites are thought to be present in the complement-like repeats that occur in clusters in all members of this receptor family. We have expressed two such complement-like repeats, CR3 and CR8, from an alpha2-macroglobulin-proteinase ligand binding region of LRP, as well as repeat 1 from the LDL receptor and examined the metal binding properties and resulting structural changes of these three repeats using changes in tryptophan and terbium fluorescence and perturbation of [1H-15N]-HSQC NMR spectra of the 15N-labeled domains from LRP. We found that all three domains contain a tight calcium binding site at physiological pH and that calcium binding results in a major structural rigidification. Changes in tryptophan fluorescence and tryptophan-sensitized terbium fluorescence indicate that the calcium binding sites are located in homologous regions in all of the repeats. Differences in the details of the perturbations, as well as in the pH dependence of calcium binding, show, however, that each metal site is distinct.
Dolmer
K
K
Department of Biochemistry and Molecular Biology, College of Medicine, University of Illinois at Chicago, Chicago, Illinois 60612-4316, USA.
Huang
W
W
Gettins
P G
PG
D000595:Amino Acid Sequence; D002118:Calcium / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000737:chemistry*; D003165:Complement System Proteins / Q000737:chemistry*; D005453:Fluorescence; D005682:Gadolinium / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics; D011971:Receptors, Immunologic / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011973:Receptors, LDL / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry; D020449:Repetitive Sequences, Amino Acid; D013725:Terbium / Q000378:metabolism; D014364:Tryptophan / Q000737:chemistry
D002135:Calcium-Binding Proteins; D026503:Low Density Lipoprotein Receptor-Related Protein-1; D010446:Peptide Fragments; D011971:Receptors, Immunologic; D011973:Receptors, LDL; D011994:Recombinant Proteins; D013725:Terbium; D014364:Tryptophan; D003165:Complement System Proteins; D005682:Gadolinium; D002118:Calcium