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H Krude

Publications and source records attributed to H Krude.

27 records · Page 2Linked to original sources
PubMed1996

The gene for the thyrotropin receptor (TSHR) as a candidate gene for congenital hypothyroidism with thyroid dysgenesis.

According to the central role of the TSH receptor for thyroid function and growth the gene for the TSH receptor is a possible candidate gene for mutations which result in an impairment of thyroid growth and function (Vassart and Dumont 1992). First evidence for the role of TSH receptor defects in the pathogenesis of congenital thyroid disorders was elucidated by the presence of activating germline mutations leading to congenital hyperthyroidism (Duprez et al., 1994). After the finding of partial loss-of-function mutations leading to hyperthyrotropinemia (Sunthornthepvarakul et al., 1995) it was speculated that a more severe phenotype with hypothyroidism and hypoplasia of the gland (thyroid dysgenesis) would be the result, if complete loss-of-function mutations like the isoleucine167 to asparagine mutation would occur in a homozygote or compound heterozygote state. The screening of TSHR gene mutations by SSCP in a well defined cohort of 100 children with congenital hypothyroidism (CH), diagnosed and followed since 1978 in the Childrens Hospital of Berlin, revealed one patient with hypoplasia of the thyroid to be positive for two compound heterozygote inactivating mutations of the TSHR gene, indicating thereby that the clinical approach to define phenotypes of interest could be helpful to understand the fundamental process of thyroid development.

Animals↗
PubMed1996

Screening for mutations of the human thyroid peroxidase gene in patients with congenital hypothyroidism.

While congenital hypothyroidism in 80-90% of the affected individuals is caused by thyroid dysgenesis (athyrosis, ectopy or hypoplasia), hypothyroidism in patients with a thyroid gland of normal position and size can be due to regulatory or enzymatic defects of thyroid hormone biosynthesis. Beside defects of thyroglobulinsynthesis, defects of the sodium-iodide-transporter or the TSH-receptor, a defect of the thyroidperoxidase, the key-enzyme of thyroid hormone biosynthesis, can cause a total iodide organification defect and thereby congenital hypothyroidism. We screened 14 of 103 patients (13.6%) with non familial congenital hypothyroidism and a normally developed thyroid gland detected by the newborn screening program with the PCR-SSCP (single-stranded-conformational-polymorphism) technique for mutations in the exons 2, 8, 9, 10 and 14 of the human thyroperoxidase gene, and in which mutations had been described previously in Dutch and Brazilian families with total organification defects. Most of the previously reported mutations were found in exons 8, 9 and 10 which code for the caralytic part of the enzyme. In two patients a GGCC-duplication in exon 8 was detected leading to a premature stop codon in exon 9. While one patient without neonatal goiter was homozygous for this mutation, the second patient was only heterozygous thus demanding another mutation on the second TPO-allel to explain the phenotype. Since the GGCC duplication is easily demonstrable by a NaeI digestion, because it creates a restriction site for this enzyme, screening for this mutation is indicated since it is easy to perform. In contrast to the perchlorate discharge test molecular genetic studies are less invasive, but as useful in making a definitive diagnosis in the individual patient. Furthermore it is the first feasible step to study the etiology and epidemiology of the so far only putative defects of thyroid hormone biosynthesis leading to congenital hypothyroidism.

Congenital Hypothyroidism↗
PubMed1992

[The human chorionic gonadotropin (hCG) receptor: a new class within the family of GTP protein coupled receptors. Epitope mapping of receptor-bound agonistic and antagonistic forms of hCG].

Human chorionic gonadotropin (hCG) is a member of the glycoprotein hormone family. It is composed of an alpha and a beta subunit, the latter being closely homologous to that of human luteinizing hormone (hLH). HCG and hLH bind to the same receptor. The molecular mass of hCG is 38 kD, up to 30% of which is contributed by the carbohydrate moieties linked to each of the two subunits. Structure-function relationship studies have indicated that both subunits interact with the receptor and that altogether four different peptide domains (two on each subunit) are responsible for high-affinity receptor binding. The carbohydrate units are responsible for expression of agonist activity: deglycosylated hCG (degly-hCG) is unable to induce a biological response (cAMP increase), despite high-affinity binding. We have previously mapped the antigenic surface of hCG and described 14 different epitopes that can be recognized by specific monoclonal antibodies (MCA), i.e. five epitopes on the alpha subunit, five on the beta subunit and four epitopes which are formed through association of the two subunits (conformational or alpha beta epitopes). The number and topography of epitopes was found to be the same on degly-hCG. This framework of immunological coordinates was then applied to probe the orientation of hCG, as well as that of the competitive antagonist degly-hCG in their receptor-bound states. With a receptor-hormone-125I-MCA sandwich approach we found that while hCG still presented two of the 14 epitopes, i.e. beta 3 and beta 5, no epitope was accessible on receptor-bound degly-hCG. This differential pattern thus correlates with the signal transduction-competence of the respective ligand. Overall, these data indicate that most of the surface of hCG is masked by moieties of the receptor. Accordingly, that moiety must itself be folded in a way that it provides a large contact surface. No portion other than the 341-residue long extracellular domain would seem capable of doing that. Computer-assisted secondary structure predictions support this view. This domain is the novel feature that distinguishes glycoprotein-hormone receptors from the other members of the G protein-coupled receptor superfamily.

Amino Acid Sequence↗
PubMed1991

Epitope mapping of the receptor-bound agonistic form of human chorionic gonadotropin (hCG) in comparison to the antagonistic form (deglycosylated hCG).

On the surface of free human chorionic gonadotropin (hCG), we can distinguish with our panel of monoclonal antibodies (MCA) 14 topographically distinct epitopes (designated alpha 1 - alpha 5, beta 1 - beta 5, alpha beta 1 - alpha beta 4, depending on the subunit they are attached to). Only 2, i.e. the adjacent beta 3 and beta 5 epitopes, of these 14 are accessible to 125I-labeled MCA binding, when hCG is first allowed to bind to the rat testis hCG receptor. This result indicates that the agonist hCG assumes a defined orientation in its receptor-bound state and that, except for that small area comprising the beta 3 and beta 5 epitopes, most of its surface is masked by the hCG receptor. We therefore asked whether the competitive antagonist deglycosylated hCG (degly-hCG), which, when free, is antigenically (as to number and topography of epitopes) indistinguishable from native hCG, would interact with the receptor differently, that is, in a way that can be discerned by this epitope accessibility paradigm. Here we describe that on receptor-bound degly-hCG the beta 3 and beta 5 epitopes were concealed as were all other epitopes. This observation, together with finding the receptor affinity of degly-hCG to be 4 times higher than that of native hCG, suggests that degly-hCG assumes a signal transduction-incompetent ligand orientation and at the same time interacts with the receptor more intensively, i.e. establishes additional ("antagonist accessory") protein-protein contacts besides those involved in agonist binding. It thus appears that the carbohydrate moieties function to prevent formation of such accessory contacts.

Animals↗
PubMed1991

Twelve of fourteen surface epitopes of receptor-bound human chorionic gonadotropin (hCG) being antibody-inaccessible suggest an extensive involvement of the long extracellular domain of the hCG receptor.

On the surface of the free (receptor-unbound) form of hCG, we have previously identified 14 topographically distinct epitopes (Schwarz et al. (1986) Endocrinology 118, 189-197; Berger et al. (1990) J. Endocrinol. 125, 301-309). Here we report that only two of them, i.e. the (adjacent) beta 3 and beta 5 epitopes, can be recognized by 125iodine-labeled monoclonal antibodies when hCG was specifically bound to the rat testis hCG receptor. The exclusive accessibility of precisely these two surface epitopes indicates that hCG assumes a defined rather than a stochastic orientation in its receptor-bound state. The inaccessibility of 12 of 14 epitopes is consistent with the idea that the 341 residues long extracellular domain of the recently cloned hCG receptor (MacFarland et al. (1989) Science 245, 494-499) is the ligand binding domain. It is proposed that the extracellular domain is folded in a way that a cavity is formed large enough to accommodate hCG. Thereby, a considerable portion of the total surface of hCG is covered, as reflected by the masking of most of its epitopes.

Animals↗
PubMed1991

Number and topography of epitopes of human chorionic gonadotropin (hCG) are shared by desialylated and deglycosylated hCG.

A previously established map of the surface epitopes of human chorionic gonadotropin (hCG) served as template for the present study in which we investigated the antigenic surfaces of two glycosylation variants of hCG, i.e. desialylated hCG (asialo-hCG) and deglycosylated hCG (degly-hCG). This map allocates five epitopes to the alpha subunit, five to the beta subunit and four alpha beta epitopes to structures formed only by the alpha/beta heterodimer holo-hCG (Schwarz et al. (1986) Endocrinology 118, 189-197; Berger et al. (1990) J. Endocrinol. 125, 301-309). Here it is described that both variants complied with this template: each of the 14 distinct monoclonal antibodies with which the epitopes of hCG were defined reacted with radiolabeled asialo-hCG and degly-hCG as well and generally bound degly-hCG with greater affinity than hCG. Moreover, every combination of capture and radiolabeled detection antibody that was either compatible or incompatible on unlabeled hCG was so also on unlabeled asialo-hCG and degly-hCG. It thus appears that alterations of the carbohydrate structure of hCG can be associated with a change in affinity between some antibodies and their respective epitopes but not with a loss of an epitope or with a change in the topographical relationships of the 14 epitopes.

Antibodies, Monoclonal↗
PubMed1991

Relationship of orientation with affinity and activity of receptor-bound glycosylation variants of human chorionic gonadotropin (hCG), as visualized by monoclonal antibodies (MCA).

When hCG was receptor-bound, only 2 epitopes (i.e. beta 3 and beta 5) out of its previously mapped total of 14 surface epitopes (10, 11, 12) could be detected by the respective 125I-MCA. Clearly, this indicates a non-random orientation of hCG in this particular state (1). Now we report that on receptor-bound desialylated (asialo-hCG) as well as on receptor-bound deglycosylated hCG (degly-hCG), the beta 3 and beta 5 epitopes were inaccessible for 125I-MCA as were the remaining epitopes, although both variants, when not receptor-bound, were indistinguishable from native hCG with respect to number and topography of epitopes. Thus, the carbohydrate (CHO) units of hCG neither seem to be part of these 14 antigenic sites nor to contribute to the affinity of receptor binding: both variants had even higher affinities than native hCG. However, since the CHO are known to be obligatory for triggering the postreceptor responses of hCG-stimulated target cells, as seen by the 50% reduced or totally abolished biological potency of asialo-hCG and degly-hCG, respectively, the here demonstrated clear-cut differences in epitope accessibility can be related to differences in receptor-bound orientations which reflect signal transduction-competent and incompetent modes of interaction with the receptor. We believe that the CHO, while not contributing to receptor binding per se, function to assure correct positioning of hCG in the ligand recognition domain to allow for proper protein-protein interactions between ligand and receptor and thus optimal activation and hence transduction of the external signal to the cell's interior. In addition, the fact that most of the surface epitopes were masked on receptor-bound hCG, represents the first experimental support for the sofar unproven hypothesis that this unusually long (i.e. 341 amino acid residues) extracellular N' terminal domain of the recently cloned hCG receptor (hCG-R) (23,24), is indeed involved in ligand recognition.

Animals↗
PubMedDate not supplied

Implications of proopiomelanocortin (POMC) mutations in humans: the POMC deficiency syndrome.

The recent discovery of the contribution of proopiomelanocortin (POMC)-derived peptides to the regulation of energy homeostasis and exocrine gland secretion in mice aroused new interest in the complex function of the endocrine POMC network. In addition, the first mutations in the gene encoding POMC have been identified in two patients affected by adrenal insufficiency, early onset severe obesity and red hair pigmentation. Therefore, the focus of this brief review will be the detailed discussion of the implications of these new findings in the physiology of the human POMC ligand-receptor system.

Amino Acid Sequence↗