Structural characterization of the ADAM 16 disintegrin loop active site.
ADAM's have various roles in intercellular adhesion and are thought to function by binding integrins through a 13 amino acid motif called the disintegrin loop. Xenopus laevis sperm express the protein ADAM 16, and peptides with the sequence of its disintegrin loop cause downstream events in eggs that require a rise in intracellular calcium similar to that occurring at fertilization. We characterized the portion of the ADAM 16 disintegrin loop responsible for causing egg activation. A peptide based on the C-terminal half of the motif, which includes a known integrin-binding sequence, is a partial agonist of calcium release. A peptide with the N-terminal sequence of the motif activates eggs in a manner virtually identical to the full-length peptide but lacks a recognized integrin-binding sequence. None of these peptides alter the permeability or fluidity of liposomes made from membrane lipids of X. laevis eggs. This result reflects the fact that the peptides do not cause calcium to leak across the egg membrane and indirectly provides evidence that they act through a receptor on the egg surface. The infrared spectrum of the full-length peptide has a strong absorption peak corresponding to a beta-turn. We predict this structure occurs at the N-terminal sequence MPKT. A rearranged peptide lacking any turns fails to activate eggs. These results provide the first structural information about the active site of an ADAM disintegrin loop. We interpret these results in terms of active site sequences from other ADAM's and the role of integrins during fertilization.
Norris
Jeffrey W
JW
Center For Biostabilization, University of California, 1 Shields Avenue, Davis, California 95616, USA. jwnorris@ucdavis.edu
Tomczak
Melanie M
MM
Oliver
Ann E
AE
Tsvetkova
Nelly M
NM
Crowe
John H
JH
Tablin
Fern
F
Nuccitelli
Richard
R
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002463:Cell Membrane Permeability / Q000187:drug effects; D003545:Cysteine / Q000737:chemistry / Q000378:metabolism; D019483:Disintegrins / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D005260:Female; D006801:Humans; D008297:Male; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D010063:Ovum / Q000737:chemistry / Q000187:drug effects; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000737:chemistry / Q000494:pharmacology; D017433:Protein Structure, Secondary; D017550:Spectroscopy, Fourier Transform Infrared; D013094:Spermatozoa / Q000737:chemistry / Q000378:metabolism; D029867:Xenopus Proteins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D014982:Xenopus laevis
D019483:Disintegrins; D008562:Membrane Glycoproteins; D010446:Peptide Fragments; D029867:Xenopus Proteins; D003545:Cysteine