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Total synthesis of the ?-subunit of human glycoprotein hormones: toward fully synthetic homogeneous human follicle-stimulating hormone.

ABSTRACT: Described herein is the first total chemical synthesis of the unique ?-subunit of the human glycoprotein hormone (?-hGPH). Unlike the biologically derived glycoprotein hormones, which are isolated as highly complex mixtures of glycoforms, ?-hGPH obtained by chemical synthesis contains discrete homogeneous glycoforms. Two such systems have been prepared. One contains the disaccharide chitobiose at the natural N-glycosylation sites. The other contains dodecamer oligosaccharides at these same sites. The dodecamer sugar is a consensus sequence incorporating the key features associated with human glycoproteins.

SUBMITTER: Aussedat B

PROVIDER: S-EPMC3288947 | biostudies-literature | 2012 Feb

REPOSITORIES: biostudies-literature

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Total synthesis of the α-subunit of human glycoprotein hormones: toward fully synthetic homogeneous human follicle-stimulating hormone.

Aussedat Baptiste B Fasching Bernhard B Johnston Eric E Sane Neeraj N Nagorny Pavel P Danishefsky Samuel J SJ

Journal of the American Chemical Society 20120206 7

Described herein is the first total chemical synthesis of the unique α-subunit of the human glycoprotein hormone (α-hGPH). Unlike the biologically derived glycoprotein hormones, which are isolated as highly complex mixtures of glycoforms, α-hGPH obtained by chemical synthesis contains discrete homogeneous glycoforms. Two such systems have been prepared. One contains the disaccharide chitobiose at the natural N-glycosylation sites. The other contains dodecamer oligosaccharides at these same sites ...[more]

PMID: 22280541

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Probing the frontiers of glycoprotein synthesis: the fully elaborated β-subunit of the human follicle-stimulating hormone.

Project description: Not available

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Two Synthetic Peptides Corresponding to the Human Follicle-Stimulating Hormone β-Subunit Promoted Reproductive Functions in Mice.

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Molecular cloning of pituitary glycoprotein alpha-subunit and follicle stimulating hormone and chorionic gonadotropin beta-subunits from New World squirrel monkey and owl monkey.

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| S-EPMC2277479 | biostudies-literature

Hypo-glycosylated human follicle-stimulating hormone (hFSH(21/18)) is much more active in vitro than fully-glycosylated hFSH (hFSH(24)).

Project description:Hypo-glycosylated hFSH(21/18) (possesses FSH?(21) and FSH?(18)bands) was isolated from hLH preparations by immunoaffinity chromatography followed by gel filtration. Fully-glycosylated hFSH(24) was prepared by combining the fully-glycosylated FSH?(24) variant with hCG? and isolating the heterodimer. The hFSH(21/18) glycoform preparation was significantly smaller than the hFSH(24) preparation and possessed 60% oligomannose glycans, which is unusual for hFSH. Hypo-glycosylated hFSH(21/18) was 9- to 26-fold more active than fully-glycosylated hFSH(24) in FSH radioligand assays. Significantly greater binding of (125)I-hFSH(21/18) tracer than hFSH(24) tracer was observed in all competitive binding assays. In addition, higher binding of hFSH(21/18) was noted in association and saturation binding assays, in which twice as much hFSH(21/18) was bound as hFSH(24). This suggests that more ligand binding sites are available to hFSH(21/18) in FSHR than to hFSH(24). Hypo-glycosylated hFSH(21/18) also bound rat FSHRs more rapidly, exhibiting almost no lag in binding, whereas hFSH(24) specific binding proceeded very slowly for almost the first hour of incubation.

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Follicle-Stimulating Hormone Glycobiology.

Project description:FSH glycosylation varies in two functionally important aspects: microheterogeneity, resulting from oligosaccharide structure variation, and macroheterogeneity, arising from partial FSHβ subunit glycosylation. Although advances in mass spectrometry permit extensive characterization of FSH glycan populations, microheterogeneity remains difficult to illustrate, and comparisons between different studies are challenging because no standard format exists for rendering oligosaccharide structures. FSH microheterogeneity is illustrated using a consistent glycan diagram format to illustrate the large array of structures associated with one hormone. This is extended to commercially available recombinant FSH preparations, which exhibit greatly reduced microheterogeneity at three of four glycosylation sites. Macroheterogeneity is demonstrated by electrophoretic mobility shifts due to the absence of FSHβ glycans that can be assessed by Western blotting of immunopurified FSH. Initially, macroheterogeneity was hoped to matter more than microheterogeneity. However, it now appears that both forms of carbohydrate heterogeneity have to be taken into consideration. FSH glycosylation can reduce its apparent affinity for its cognate receptor by delaying initial interaction with the receptor and limiting access to all of the available binding sites. This is followed by impaired cellular signaling responses that may be related to reduced receptor occupancy or biased signaling. To resolve these alternatives, well-characterized FSH glycoform preparations are necessary.

| S-EPMC6534497 | biostudies-literature

Project description:Synthesis of the FSH beta-subunit (FSHbeta) is critical for normal reproduction in mammals, and its expression within the pituitary gonadotrope is tightly regulated by activin. Here we show that Runt-related (RUNX) proteins, transcriptional regulators known to interact with TGFbeta signaling pathways, suppress activin induction of FSHbeta gene expression. Runx2 is expressed within the murine pituitary gland and dramatically represses activin-induced FSHbeta promoter activity, without affecting basal expression in LbetaT2 cells, an immortalized mouse gonadotrope cell line. This repressive effect is specific, because RUNX2 induces LHbeta transcription (with or without activin) and does not interfere with GnRH induction of either gonadotropin beta-subunit gene. Analysis of the murine FSHbeta promoter by transfection and gel shift assays reveals that RUNX2 repression localizes to a Runx-binding element at -159/-153, which is adjacent to a previously recognized region critical for activin induction. Mutation of this -153 activin-response element or, indeed, any of the five activin-responsive regions prevents activin induction and, in fact, RUNX2 suppression, instead converting RUNX2 to an activator of the FSHbeta gene. Although the Runx-binding element is required for RUNX2-mediated repression of FSHbeta induction by either activin or Smad3, confirming a functional role of this novel site, protein interactions in addition to those between RUNX2 and Smads are necessary to account for full repression of activin induction. In summary, the present study provides evidence for Runx2-mediated repression of activin-induced FSHbeta gene expression and reveals the context dependence of Runx2 action in hormonal regulation of the gonadotropin genes.

| S-EPMC2875819 | biostudies-literature

Periodate oxidation studies of human pituitary follicle-stimulating hormone.

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| S-EPMC1185093 | biostudies-other

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