Biochemical characterization of human eosinophil Charcot-Leyden crystal protein (lysophospholipase).
Lysophospholipase from human eosinophils is a protein previously considered based upon antigenic, enzymatic, and electrophoretic similarities to be the single component of Charcot-Leyden crystals, which are formed in vivo in association with eosinophilic diseases. The identity of eosinophil lysophospholipase and solubilized Charcot-Leyden crystal protein is now established by biochemical criteria, and a basis for the ease of aggregation and crystallization of the protein is identified in its prominent hydrophobicity. Chromatographically purified enzyme and Charcot-Leyden crystal protein formed in vitro functioned as lysophospholipases with identical Michaelis constants (Km approximately equal to 22 microM) for the substrate lysopalmitoylphosphatidylcholine and had blocked amino-terminal residues and almost identical amino acid compositions. The propensity of lysophospholipase to aggregate was not due to extensive intermolecular disulfide bonding because it contained a single cysteine residue as assessed by amino acid analyses and incorporated 0.986 mol of p-chloromercuribenzoic acid/mol of native enzyme or 0.958 mol of iodoacetic acid/mol of reduced and denatured enzyme. By equilibrium dialysis, lysophospholipase bound 3.820 g of detergent/g of protein in 1% sodium dodecyl sulfate and 0.506 g of detergent/g of protein in 10 mM sodium deoxycholate. In addition, monomeric protein demonstrated enhanced binding of detergent as evidenced by its aberrantly rapid electrophoretic mobility in 1%, but not 0.1%, sodium dodecyl sulfate. The hydrophobic nature of this protein, which accounts for 10% of the protein of the eosinophil, may contribute to its unique propensity for crystallization in vivo.
D000596:Amino Acids / Q000032:analysis; D002729:Chloromercuribenzoates / Q000494:pharmacology; D003460:Crystallization; D004804:Eosinophils / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D008245:Lysophospholipase / Q000097:blood* / Q000302:isolation & purification; D008970:Molecular Weight; D010740:Phospholipases / Q000097:blood*; D020245:p-Chloromercuribenzoic Acid
D000596:Amino Acids; D002729:Chloromercuribenzoates; D020245:p-Chloromercuribenzoic Acid; D010740:Phospholipases; D008245:Lysophospholipase