Purification and characterization of proteoliaisin, a coordinating protein in fertilization envelope assembly.
We report the purification and characterization of proteoliaisin, a protein that participates in the assembly of the sea urchin fertilization envelope. Proteoliaisin was purified from egg cortical granule exudate to greater than 99% homogeneity using chromatography on DEAE-Sepharose and on phenyl-Sepharose. Native proteoliaisin is a highly asymmetric protein (f/fo = 2.0) composed of a single Mr approximately 230,000 peptide. Its asymmetry was demonstrated both by analytical ultracentrifugation and by nondenaturing polyacrylamide gel electrophoresis, a novel analysis that detects molecular asymmetry in heterogeneous protein mixtures. Proteoliaisin is enriched in six amino acids: aspartic acid/asparagine, glutamic acid/glutamine, glycine, and cysteine, which account for over 50% of its mass. Nearly all of the cysteine residues are disulfide bonded. The protein contains a small proportion of aromatic amino acids with phenylalanine greater than tyrosine greater than tryptophan. At neutral pH its absorbance maximum is at 274.5 nm, with an extinction coefficient of 0.43 ml mg-1 cm-1. Proteoliaisin forms a 1:1 Ca2+-stabilized complex with ovoperoxidase, another component of the fertilization envelope, with Kd = 1.1 X 10(-6) M. Proteoliaisin, a constituent of the specialized echinoderm extracellular matrix called the fertilization envelope, has certain structural similarities to mammalian extracellular matrix proteins.
Weidman
P J
PJ
Department of Biochemistry, University of Washington, Seattle 98195.
Teller
D C
DC
Shapiro
B M
BM
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D005306:Fertilization; D006023:Glycoproteins / Q000302:isolation & purification*; D008562:Membrane Glycoproteins / Q000302:isolation & purification*; D008970:Molecular Weight; D010063:Ovum / Q000032:analysis; D011487:Protein Conformation; D012617:Sea Urchins; D014461:Ultracentrifugation
D000596:Amino Acids; D006023:Glycoproteins; D008562:Membrane Glycoproteins; C044807:proteoliaisin protein, Strongylocentrotus purpuratus