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Glycosylation Effects on FSH-FSHR Interaction Dynamics: A Case Study of Different FSH Glycoforms by Molecular Dynamics Simulations.

Meher BR, Dixit A, Bousfield GR, Lushington GH - PLoS ONE (2015)

Bottom Line: However, substantial qualitative differences emerge between FSH15 and FSH24 when FSH is decorated with a much larger, tetra-antennary glycan.Specifically, the FSHR complex with hypo-glycosylated FSH15 is observed to undergo a significant conformational shift after 5-10 ns of simulation, indicating that FSH15 has greater conformational flexibility than FSH24 which may explain the more favorable FSH15 kinetic profile.FSH15 also exhibits a stronger binding free energy, due in large part to formation of closer and more persistent salt-bridges with FSHR.

View Article: PubMed Central - PubMed

Affiliation: Bioinformatics Core Facility, University of Kansas, Lawrence, Kansas, United States of America.

ABSTRACT
The gonadotropin known as follicle-stimulating hormone (FSH) plays a key role in regulating reproductive processes. Physiologically active FSH is a glycoprotein that can accommodate glycans on up to four asparagine residues, including two sites in the FSHα subunit that are critical for biochemical function, plus two sites in the β subunit, whose differential glycosylation states appear to correspond to physiologically distinct functions. Some degree of FSHβ hypo-glycosylation seems to confer advantages toward reproductive fertility of child-bearing females. In order to identify possible mechanistic underpinnings for this physiological difference we have pursued computationally intensive molecular dynamics simulations on complexes between the high affinity site of the gonadal FSH receptor (FSHR) and several FSH glycoforms including fully-glycosylated (FSH24), hypo-glycosylated (e.g., FSH15), and completely deglycosylated FSH (dgFSH). These simulations suggest that deviations in FSH/FSHR binding profile as a function of glycosylation state are modest when FSH is adorned with only small glycans, such as single N-acetylglucosamine residues. However, substantial qualitative differences emerge between FSH15 and FSH24 when FSH is decorated with a much larger, tetra-antennary glycan. Specifically, the FSHR complex with hypo-glycosylated FSH15 is observed to undergo a significant conformational shift after 5-10 ns of simulation, indicating that FSH15 has greater conformational flexibility than FSH24 which may explain the more favorable FSH15 kinetic profile. FSH15 also exhibits a stronger binding free energy, due in large part to formation of closer and more persistent salt-bridges with FSHR.

No MeSH data available.


FSH glycoforms result from partial glycosylation of the hFSHβ subunit.The variants are defined by the molecular weights of the FSHβ subunit based on Western blotting experiments. The primary structures are indicated as solid lines, 92 residues for FSHα and 111 for FSHβ. The tuning forks represent N-glycans, when present. The N is the single-letter code for asparagine and the superscript represents each residue’s position in the primary structure.
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pone.0137897.g002: FSH glycoforms result from partial glycosylation of the hFSHβ subunit.The variants are defined by the molecular weights of the FSHβ subunit based on Western blotting experiments. The primary structures are indicated as solid lines, 92 residues for FSHα and 111 for FSHβ. The tuning forks represent N-glycans, when present. The N is the single-letter code for asparagine and the superscript represents each residue’s position in the primary structure.

Mentions: Evidence suggests that full glycosylation of FSHα is consistently required for normal biological functions [5,6] but indications are that FSHβ exhibits biological functionality (albeit with varying efficacy toward different cellular targets) when glycosylated at one or both FSHβ N-glycosylation sites [7,12,13]. For the purposes of this study, isoform nomenclature relates to approximate FSHβ molecular weight determined in Western blots, which (with allowances for variation per precise glycan formulation) can be summarized as: FSH24 refers to fully glycosylated FSH (i.e., the heterodimer with glycans at both FSHβ asparagine residues; molecular weight approximately 24 KDa), FSH15 for fully hypo-glycosylated FSHβ without substitutions at either asparagine (MW ≈ 15 KDa), and FSH21 for hypo-glycosylated FSHβ with substitution at only Asn7 (MW ≈ 21 KDa) and FSH18 for hypo-glycosylated FSH with substitution at only Asn24 (MW ≈ 18 KDa). The FSHα subunit always possesses both N-glycans and migrates as a single band following SDS-PAGE, albeit with varying mobilities depending on the glycan populations attached. These FSH glycoform variants are summarized in Fig 2. The composition and structure of the tetra-antennary glycan used in the simulations is shown in schematic form in Fig 3 using the Oxford Glycobiology Institute system [14,15], with one modification in order to provide the readers a better understanding of the structure—the α/β 1–6 linkage line is made longer indicating the carbon atom is exocyclic. All other linkages involve ring carbons.


Glycosylation Effects on FSH-FSHR Interaction Dynamics: A Case Study of Different FSH Glycoforms by Molecular Dynamics Simulations.

Meher BR, Dixit A, Bousfield GR, Lushington GH - PLoS ONE (2015)

FSH glycoforms result from partial glycosylation of the hFSHβ subunit.The variants are defined by the molecular weights of the FSHβ subunit based on Western blotting experiments. The primary structures are indicated as solid lines, 92 residues for FSHα and 111 for FSHβ. The tuning forks represent N-glycans, when present. The N is the single-letter code for asparagine and the superscript represents each residue’s position in the primary structure.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4581761&req=5

pone.0137897.g002: FSH glycoforms result from partial glycosylation of the hFSHβ subunit.The variants are defined by the molecular weights of the FSHβ subunit based on Western blotting experiments. The primary structures are indicated as solid lines, 92 residues for FSHα and 111 for FSHβ. The tuning forks represent N-glycans, when present. The N is the single-letter code for asparagine and the superscript represents each residue’s position in the primary structure.
Mentions: Evidence suggests that full glycosylation of FSHα is consistently required for normal biological functions [5,6] but indications are that FSHβ exhibits biological functionality (albeit with varying efficacy toward different cellular targets) when glycosylated at one or both FSHβ N-glycosylation sites [7,12,13]. For the purposes of this study, isoform nomenclature relates to approximate FSHβ molecular weight determined in Western blots, which (with allowances for variation per precise glycan formulation) can be summarized as: FSH24 refers to fully glycosylated FSH (i.e., the heterodimer with glycans at both FSHβ asparagine residues; molecular weight approximately 24 KDa), FSH15 for fully hypo-glycosylated FSHβ without substitutions at either asparagine (MW ≈ 15 KDa), and FSH21 for hypo-glycosylated FSHβ with substitution at only Asn7 (MW ≈ 21 KDa) and FSH18 for hypo-glycosylated FSH with substitution at only Asn24 (MW ≈ 18 KDa). The FSHα subunit always possesses both N-glycans and migrates as a single band following SDS-PAGE, albeit with varying mobilities depending on the glycan populations attached. These FSH glycoform variants are summarized in Fig 2. The composition and structure of the tetra-antennary glycan used in the simulations is shown in schematic form in Fig 3 using the Oxford Glycobiology Institute system [14,15], with one modification in order to provide the readers a better understanding of the structure—the α/β 1–6 linkage line is made longer indicating the carbon atom is exocyclic. All other linkages involve ring carbons.

Bottom Line: However, substantial qualitative differences emerge between FSH15 and FSH24 when FSH is decorated with a much larger, tetra-antennary glycan.Specifically, the FSHR complex with hypo-glycosylated FSH15 is observed to undergo a significant conformational shift after 5-10 ns of simulation, indicating that FSH15 has greater conformational flexibility than FSH24 which may explain the more favorable FSH15 kinetic profile.FSH15 also exhibits a stronger binding free energy, due in large part to formation of closer and more persistent salt-bridges with FSHR.

View Article: PubMed Central - PubMed

Affiliation: Bioinformatics Core Facility, University of Kansas, Lawrence, Kansas, United States of America.

ABSTRACT
The gonadotropin known as follicle-stimulating hormone (FSH) plays a key role in regulating reproductive processes. Physiologically active FSH is a glycoprotein that can accommodate glycans on up to four asparagine residues, including two sites in the FSHα subunit that are critical for biochemical function, plus two sites in the β subunit, whose differential glycosylation states appear to correspond to physiologically distinct functions. Some degree of FSHβ hypo-glycosylation seems to confer advantages toward reproductive fertility of child-bearing females. In order to identify possible mechanistic underpinnings for this physiological difference we have pursued computationally intensive molecular dynamics simulations on complexes between the high affinity site of the gonadal FSH receptor (FSHR) and several FSH glycoforms including fully-glycosylated (FSH24), hypo-glycosylated (e.g., FSH15), and completely deglycosylated FSH (dgFSH). These simulations suggest that deviations in FSH/FSHR binding profile as a function of glycosylation state are modest when FSH is adorned with only small glycans, such as single N-acetylglucosamine residues. However, substantial qualitative differences emerge between FSH15 and FSH24 when FSH is decorated with a much larger, tetra-antennary glycan. Specifically, the FSHR complex with hypo-glycosylated FSH15 is observed to undergo a significant conformational shift after 5-10 ns of simulation, indicating that FSH15 has greater conformational flexibility than FSH24 which may explain the more favorable FSH15 kinetic profile. FSH15 also exhibits a stronger binding free energy, due in large part to formation of closer and more persistent salt-bridges with FSHR.

No MeSH data available.