Amino Acid Sequence of the ,8 Subunit of Follicle-stimulating Hormone from Human Pituitary Glands*

The p subunit of follicle-stimulating hormone (FSH-@) from human pituitary glands was reduced and S-aminoethylated prior to thermolytic, tryptic, and chymotryptic digestions. Each digest was gel-filtered on Sephadex G-50 to separate the glycopeptides from the peptides. The glycopeptides and the peptides were isolated by high voltage paper electrophoresis at pH 6, 3.5, and 2.0. The purity of the isolated peptides was confirmed by amino acid analyses. The amino acid sequences of peptides were determined by Edman degradation followed by subtractive amino acid analysis and, in certain cases, confirmed by dansylation. COOH-terminal sequences of the peptides were determined by digestion with carboxypeptidases A and B and by hydrazinolysis. The tryptophan content of human follicle-stimulating hormone, of the /3 subunit of human follicle-stimulating hormone, and of the glycopeptides obtained from the enzymic digests was determined by fluorescence spectra, titration against N-bromosuccinimide, calorimetric estimation with p-dimethyl aminobenzaldehyde, hydrolysis with methane sulfonic acid containing 0.2% tryptamine followed by amino acid analysis, microbiological assay, and sequence analysis. The presence of 1 tryptophan residue in the p subunit was indicated. Human FSH-P consists of 118 amino acid residues, with a predominant proportion of molecules having 108 residues due to microheterogeneity at the NH, and COOH termini. The sequence indicates an abundance of threonine, glutamic acid, and cysteine residues. The NH,-terminal portion (up to 32 residues) shows homology with the /3 subunits of other glycoprotein hormones such as luteinizing hormone, human chorionic gonadotropin, and thyroid-stimulating hormone. However, the sequence differs a great deal from the others in the rest of the molecule, confirming the specificity of the fi subunit of these hormones. There are asparagine residues at positions 1, 7, 24, and 41; glutamine residues at positions 48, 81, and 111; and two carbohydrate moieties linked to asparagine at positions 7 and 24. The primary structure of human FSH-P reported here has sequences between residues 1 and 20, 28 and 43, and 47 and 108 similar to those reported by Shome and Parlow (Shome, B., and Parlow, A. F. (1974) J. Clin. Endocrinol. Metab. 39, 187-190), but different at positions 44 to 46, 21 to 28, and at the COOH terminus. The alignment of the cysteines in the first 32 residues of the @ subunit of human follicle-stimulating hormone (hFSH-P), luteinizing hormone (hLH-P), thyroid-stimulating hormone (hTSH-P), and chorionic gonadotropin (hCG-p) showed that the sequence of the p subunit of human follicle-stimulating hormone between alignment positions 30 and 38 is identical with the other hormones except that tryptophan occurs at position 33 in human FSH-/3 instead of isoleucine. When compared to bovine thyroidstimulating hormone-p subunit (bTSH-P) and ovine luteinizing hormone-P subunit (oLH-P), the sequence between positions 28 and 38 is identical in both hFSH-fi and bTSH-@, except that valine occurs at position 33 in bTSH-0 in place of tryptophan in hFSH-0. Human FSH-0, hCG-fi, and oLH-fl contain carbohydrate moieties at locations corresponding to alignment position 13, whereas bovine and human TSH-P and hLH-fi do not. It is noteworthy that the sequence between alignment positions 26 and 29 is Cys-Leu-Thr-Ileu in hFSH-P, hTSH-& and bTSH-P, whereas oLH-fi contains the sequence Cys-IleuThr-Phe, and hCG-@ and hLH-P contain the sequence Cys-Ileu-Thr-Val.