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Biomedical subjects

J G Pierce

Publications and source records attributed to J G Pierce.

At least 55 records · Page 3Linked to original sources
PubMed1976

In vitro activation of glycoprotein hormones. Hybridization of subunits from thyrotropin, lutropin and human choriogonadotropin.

In vitro assembly of thyrotropin alpha and beta subunits led to an increase in content of alpha helix and beta sheet very similar to that found for gonadotropins. This association-dependent active folding involved the burying of three tyrosine residues tentatively assigned to Tyr alpha 41, Tyr beta 37 and Tyr beta 59 and common to all studied glycoprotein hormones. In vitro hybridizations between alpha and beta subunits of various hormones (thyrotropin, lutropin and choriogonadotropin) from different species (ovine, bovine and human) triggered the same molecular events as assembly of homologous subunits: the burying of three tyrosine residues and the increase of periodic structure of the folding. These changes are slow, time-dependent processes. Rates and yields of hybrid formation measured by sedimentation analysis and difference spectroscopy of tyrosines are identical, within experimental error, with the rates and yields measured by the recovery of the biological activity either the stimulation of chick thyroids for thyrotropin-beta hybrids or binding to porcine testis receptors for gonadotropin-beta hybrids. Whatever the origin of the alpha subunit, the thyrotropin-beta hybrids were not able to bind to testis receptors although active on chick thyroids. Rates and yields of hybrid formation essentially depended on the origin of the beta subunit. All the hybrids could be dissociated at acid pH with rates similar to those of native hormone. The extension to thyrotropin and various hybrids of the structural features of the in vitro assembly already recognized for gonadotropins strengthens the hypothesis that one deals with a basic activation process which also occurs in vivo after the synthesis of the subunits.

Animals↗
PubMed1976

Structure and structure-function relationships in glycoprotein hormones.

Relationships between the sequences of thyrotropin (thyroid-stimulating hormone, TSH), lutropin (luteinizing hormone, LH), human choriogonadotropin (chorionic gonadotropin, hCG) and follitropin (follicle-stimulating hormone, FSH) are now well established. Each beta-subunit contains six disulphide bonds and considerable homology is seen when all four linear sequences are aligned with half-cystine residues in juxtaposition. Major questions about the tertiary structures of the subunits and their interactions to form active hormone remain. Determination of the disulphide bridges in both alpha- and beta-subunits has not yielded to usual methods and conflicting data about the alpha-subunit have been reported. Partial reduction of the beta-subunits of LH and TSH with subsequent labelling of the cysteines formed has shown that a single bond is first reduced. This bond is between positions 93 and 100 in LH-beta and the corresponding positions 88-95 in TSH-beta. Thus, as would be expected from the fact that interhormone hybrids can be made with the common alpha-subunits, the chemical data, though still limited, indicate similar tertiary structures for the different beta-subunits. To investigate whether other useful intermediates can be obtained after partial reduction, we have studied reduction and derivative formation in various conditions. Intact LH is more resistant to reduction than either its alpha- or beta-subunit but no intermediates have been observed which are not present after partial reduction of individual subunits. Preliminary experiments on the reoxidation of fully reduced alpha-subunit show that the reoxidized material will recombine with native beta-subunits to yield biologically active TSH or LH. Studies from this and other laboratories on chemical modifications of several amino acid residues of glycoprotein hormones and their subunits are also summarized.

Amino Acid Sequence↗
PubMed1975

The carboxylic acid groups of bovine luteinizing hormone. The effects of their modification on receptor site binding and subunit-subunit interaction.

The modification of the carboxyl groups of the subunits of bovine luteinizing hormone to neutral derivatives by carbodiimide-mediated coupling with glycine methyl ester has been studied. The modified alpha subunit, which has 8 residues of glycine methyl ester incorporated, will no longer recombine with native beta (hormone-specific) subunit, but the modified beta subunit, with 6 to 7 glycine methyl esters incorporated, will recombine with native alpha to yield a partially active hormone. Derivatization of the intact hormone results in dissociation to subunits together with formation of a major side product which is covalently cross-linked. Significant cross-linked product was not obtained during modification of individual subunits, thus indicating an orientation between an activated carboxyl group(s) and a nucleophile(s) in the intact hormone which favors coupling. Separation of subunits from the derivatized, noncross-linked fraction by countercurrent distribution reveals a heterogeneous preparation of the modified alpha subunit which also will not recombine with either a native or modified beta subunit. The beta subunit from the modified intact hormone was indistinguishable from the modified isolated beta subunit in amino acid composition and in ability to recombine with native alpha subunit. The results are consonant with data from this and other laboratories in which various modifications of the alpha chain, the subunit common to the glycoproteins, more seriously affect recombination than similar modifications of the beta subunits. The number of carboxyl groups modified in each subunit is compatible with but not in total agreement with assignments of amides reported from sequence studies.

Amino Acid Sequence↗
PubMed1975

Secretion of alpha subunits of luteinizing hormone (LH) by the anterior pituitary.

Free alpha subunit chains of the glycopeptide pituitary hormones have been found in the sera of normal subjects and postmenopausal women. To ascertain whether the alpha subunit of LH is directly secreted by the pituitary or formed as a result of degradation of intact LY in the periphery, alpha subunits and intact LH were measured by radioimmunoassay in human volunteers after LRF stimulation and purified LH infusion. In 4 subjects a loading dose of 90 IU, followed by the infusion of 22.5 IJ of purified human LH over 30 min, produced peak serum LH levels of 41 mIU/ml but no change in alpha subunit levels of 35 IU of purified human LH to an additional 4 subjects, produced peak LH levels of 8* mIU/ml, but again, no change in alpha subunits. In the same two groups of subjects 100 mug of LRF produced peak LH levels of 25 mIU/ml and 75 mIU/ml, respectively, with significant alpha subunit elevations at 20 min of 1.7 ng/ml and 2.7 ng/ml, respectively. In separate groups of men LRF was administered over a wide dose range of 1 to 3,000 mug and LH and the alpha subunit measured. A dose-response curve existed over the entire LRF dose range for blood LH; no minimum or maximum plateaus were observed over the range studied. However, the alpha chain response appeared to reach a maximal plateau at a dose of 100 mug of LRF. The results are compativle with the hypothesis that the alpha subunits appearing in the peripheral circulation in response to LRF are due to secretion by the anterior pituitary and not due to peripheral degradation of intact secreted LH.

Dose-Response Relationship, Drug↗
PubMed1973

Neutralizing and non-neutralizing antibodies to bovine thyroid-stimulating hormone and its subunits.

To test the possibility that the long-acting thyroid stimulator (LATS) might represent an immune complex either of thyroid-stimulating hormone (TSH) with anti-TSH or of a subunit of TSH with an appropriate antibody, we immunized rabbits with bovine TSH (bTSH), bLH (luteinizing hormone), and their alpha and beta subunits (bTSHalpha and bTSHbeta). Binding, neutralizing, and nonneutralizing antibodies were demonstrated in the antisera obtained. First, antisera to TSH, TSHbeta, and TSHalpha all bound [(125)I]TSH and [(125)I]TSHbeta. Anti-bTSHbeta antisera bound [(125)I]bTSHbeta better than did anti-TSH sera, while the binding of [(125)I]bTSH was similar with both types of antiserum. Second, the thyroid-stimulating activity (McKenzie bioassay) of TSH could be neutralized by incubation with various dilutions of anti-TSH or anti-TSHbeta. Finally, when incubation mixtures containing TSH and dilutions of anti-TSHbeta antisera that only partially neutralized TSH were treated with an antiserum against rabbit immunoglobulins to precipitate immune complexes, the bioassay response of the TSH was abolished. This phenomenon was not observed when antiserum to the intact hormone was substituted in the incubation mixture. The removal of TSH biological activity from a mixture of TSH and anti-bTSHbeta by addition of an anti-immunoglobulin indicated that biologically active immune complexes were formed between TSH and anti-TSHbeta but not between TSH and anti-TSH. The time-course of the bioactivity and several other characteristics of these complexes differentiate them from LATS.

Animals↗
PubMed1973

The effects on rabbits of immunization with bovine thyroid-stimulating hormone and its subunits.

Rabbits were immunized with bovine thyroid-stimulating hormone (bTSH), bovine Inteinizing hormone (bLH), and their subunits. In two immunization experiments, thyroid-stimulating activity was found in the serum of 6 out of 12 rabbits immunized with bTSHbeta subunits. The thyroid-stimulating activity in the anti-bTSHbeta sera was greater at 2 h than at 8, was eluted with the globulin fraction from Sephadex G-100, was completely neutralized by both anti-bTSH and anti-rabbit gamma globulin, and was completely suppressed by administration of triiodothyronine (T(3)) to the immunized rabbit. These findings led to the conclusion that the thyroid-stimulating activity resided in soluble complexes of rabbit TSH bound to anti-bTSHbeta. Two of nine rabbits immunized with bTSH developed thyroid-stimulating activity in their serum, but it was nonsuppressible by T(3). None of the animals immunized with bTSHalpha, bLH, bLHbeta, or bLHalpha developed serum thyroid-stimulating activity.Hypopituitary hypothyroidism, evidenced by decreased serum thyroxine (T(4)) and thyroidal (131)I uptake and by the histologic appearance of large follicles with flat cells, was found in the bTSHbeta- and bTSH-immunized animals, despite the presence of thyroid-stimulating activity in the serum of many. The reasons for this paradox are unclear; possibly the complexes block the effect of TSH on the rabbit thyroid.

Animals↗