A subunit interaction site in human luteinizing hormone: identification by photoaffinity cross-linking.
The sites of noncovalent association between the alpha- (common) and beta- (hormone-specific) subunits in the glycoprotein hormones [LH, human CG (hCG), FSH, and TSH] remain to be completely defined. This information is essential to efforts to map the three-dimensional structure of the hormones and to help explain the stability of the hormone heterodimer in the circulation. Among numerous approaches that have been employed to identify these sites of subunit interaction, chemical or photoaffinity cross-linking has the advantage of identifying the complementary sites of contact on the respective subunits. We have used the stable photoaffinity ligand, L-benzoylphenylalanine (Bpa), to evaluate subunit interaction by the hLH beta sequence (1-15). A peptide from this region of hCG beta has been shown previously to inhibit subunit association. The 3H-labeled Bpa was incorporated into the peptide at position 8 (Trp) during solid-phase synthesis. After incubation with alpha under long-wavelength (366 nm) ultraviolet light, a Bpa- (1-15)-labeled-alpha-fraction was isolated. Control experiments showed the binding to be covalent and specific to (1-15). Two other Bpa-labeled beta-fragments, the receptor-binding loop (38-57) and the (30-43) peptide containing the highly conserved CAGY sequence, did not cross-link. The site of contact in alpha was localized by peptide mapping and sequence analysis to the N-terminal fragment (18-33), most likely at Met-29 or Gly-30. Both the alpha- and beta-sites are adjacent to or overlapping receptor-binding regions. Subunit contact sites and receptor-binding segments may thus be oriented in close proximity to provide a multicomponent receptor-binding domain imparting full activity to the native hormone.
Keutmann
H T
HT
Department of Medicine, Massachusetts General Hospital, Boston 02114.
Rubin
D A
DA
D000345:Affinity Labels / Q000378:metabolism; D000595:Amino Acid Sequence; D001665:Binding Sites; D006063:Chorionic Gonadotropin / Q000378:metabolism*; D015292:Glycoprotein Hormones, alpha Subunit / Q000378:metabolism*; D006801:Humans; D007986:Luteinizing Hormone / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D010649:Phenylalanine / Q000031:analogs & derivatives* / Q000378:metabolism
D000345:Affinity Labels; D006063:Chorionic Gonadotropin; D015292:Glycoprotein Hormones, alpha Subunit; D010649:Phenylalanine; D007986:Luteinizing Hormone; C053764:benzoylphenylalanine