Structural studies on equine glycoprotein hormones. Amino acid sequence of equine lutropin beta-subunit.

G R Bousfield,W K Liu,H Sugino,D N Ward

doi:10.1016/s0021-9258(18)47457-1

Abstract

The amino acid sequence was determined for equine lutropin beta (eLH beta). Large fragments were derived from reduced, carboxymethylated eLH beta by digestion with Staphylococcus aureus V8 protease, by cyanogen bromide cleavage, and by cleavage of acid-labile Asp-Pro bonds. The fragments were purified by gel filtration and high performance liquid chromatography (HPLC). The fragments were sequenced by automated Edman degradation to establish the primary structure of eLH beta. Some peptides were further digested with chymotrypsin and the resulting peptides purified by HPLC. In addition to sequencing by automated Edman degradation, these were also sequenced by the complementary 5-dimethylaminonaphthalene-1-sulfonyl-Edman procedure which enabled us to directly identify glycosylated amino acids. The eLH beta subunit is a glycoprotein of 149 amino acids containing both N- and O-linked oligosaccharides. It possesses a COOH-terminal extension similar to that seen in human chorionic gonadotropin. Carboxypeptidase Y digestions suggest that the COOH terminus is blocked by glycosylation. Interestingly, the amino acid sequence of eLH beta is identical to that of equine chorionic gonadotropin beta (Sugino, H., Bousfield, G. R., Moore, W. T., and Ward, D. N. (1987)J. Biol. Chem. 262, 8603-8609).

Highlights

From The University of Texas System Cancer Center, M.D
The fragments were purified by gel filtration there is little cross-reactivity between a given glycoprotein and high performance liquid chromatography (HPLCho)r.mone and the othertwo receptors
In addition to sequencing by automatEeddman the FSH activity observed with these hormones results from degradation, thesewere sequenced by the complet-he lack of specificity of the FSH receptors of the rat [22], mentary 6-dimethylaminonaphthalene- 1-sulfonyl-Ed- and thisprobably extends to those of the pig, cow,and donkey man procedure which enabled us to directly identify as well [20,21,22,23,24]

Summary

Introduction

From The University of Texas System Cancer Center, M.D. Anderson Hospital and Tumor Institute, Department of Biochemistry and Molecular Bialogy, Houston, Texas 77030. The predicted sequence for human CG and LH) sharae threonineresidue and thisresidue the rat LHBwould have only four or five amino acids in is not glycosylated, whereas hCGp and all the otherglycopro- common and that of mouse TSHP would have none in comtein hormone 8-subunits have an asparagine residue, and an mon.

Results

Conclusion