Purification and structural characterization of the CD11b/CD18 integrin alpha subunit I domain reveals a folded conformation in solution.
The alpha subunits of the leukocyte CD11/CD18 integrins contain an approximately 200 amino acid 'inserted' or I domain. The I domain of the cell-surface Mac-1 (CD11b/CD18) integrin has been shown to be the major recognition site for several adhesion ligands, including iC3b, fibrinogen, factor X, and ICAM-1. The I domain from the Mac-1 alpha subunit has been expressed in Escherichia coli as a soluble GST-fusion protein containing a factor Xa sensitive cleavage site. Analytical characterization of the purified I domain reveals that it is obtained in very high quality at high yields. CD and NMR spectra indicate that I domain adopts a predominantly folded structure in solution, independent of the remainder of the alpha subunit. Addition of Ca2+ and Mg2+ did not significantly perturb the structural conformation.
Fairbanks
M B
MB
Upjohn Laboratories, Upjohn Company, Kalamazoo, MI 49007, USA.
Pollock
J R
JR
Prairie
M D
MD
Scahill
T A
TA
Baczynskyj
L
L
Heinrikson
R L
RL
Stockman
B J
BJ
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D018821:CD18 Antigens / Q000737:chemistry* / Q000302:isolation & purification; D002118:Calcium / Q000494:pharmacology; D002942:Circular Dichroism; D004926:Escherichia coli / Q000235:genetics; D015951:Factor Xa; D005982:Glutathione Transferase / Q000235:genetics; D007962:Leukocytes / Q000737:chemistry; D016177:Macrophage-1 Antigen / Q000737:chemistry* / Q000302:isolation & purification; D008274:Magnesium / Q000494:pharmacology; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D011487:Protein Conformation / Q000187:drug effects; D017510:Protein Folding; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D017421:Sequence Analysis
D000596:Amino Acids; D018821:CD18 Antigens; D016177:Macrophage-1 Antigen; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase; D015951:Factor Xa; D008274:Magnesium; D002118:Calcium