CD14: physical properties and identification of an exposed site that is protected by lipopolysaccharide.
Soluble CD14 (sCD14) is a 55-kDa serum protein that binds lipopolysaccharide (LPS) and mediates LPS-dependent responses in a variety of cells. Using recombinant sCD14 expressed in Chinese hamster ovary (CHO) cells, we examined the structural characteristics of sCD14 and sCD14.LPS complexes. The circular dichroism and fluorescence spectra of the sCD14 indicate that it contains substantial beta-sheet (40%) and a well-defined tertiary structure with the tryptophan residues located in environments with different degrees of hydrophobicity and solvent exposure. The spectra of the sCD14.LPS complex are identical within experimental error to the uncomplexed sCD14. Changes in surface accessibility upon LPS binding were examined using limited proteolysis with endoproteinase Asp-N. This analysis revealed that aspartic acid residues at amino acids 57, 59, and 65 are susceptible to cleavage by Asp-N, while the same residues are protected from proteolytic cleavage in the sCD14.LPS complex. These results suggest that a region including amino acids 57 to 64 is involved in LPS binding by sCD14.
McGinley
M D
MD
Amgen Inc., Amgen Center, Thousand Oaks, California 91320.
Narhi
L O
LO
Kelley
M J
MJ
Davy
E
E
Robinson
J
J
Rohde
M F
MF
Wright
S D
SD
Lichenstein
H S
HS
D000595:Amino Acid Sequence; D000818:Animals; D015703:Antigens, CD / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D015214:Antigens, Differentiation, Myelomonocytic / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D001665:Binding Sites; D016466:CHO Cells; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D006224:Cricetinae; D006801:Humans; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides / Q000302:isolation & purification / Q000378:metabolism*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D013050:Spectrometry, Fluorescence; D014162:Transfection
D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D010446:Peptide Fragments; D011994:Recombinant Proteins