Characterization and charge distribution of the asparagine-linked oligosaccharides on secreted mouse thyrotropin and free alpha-subunits
Mouse hemipituitaries in vitro secrete TSH, composed of an alpha-beta heterodimer, as well as excess (free) alpha-subunits. By dual metabolic labeling with (35S)sulfate and (3H)mannose, we have characterized oligosaccharides from secreted TSH alpha, TSH beta, and free alpha-subunits released from the apoprotein by enzymatic deglycosylation. Oligosaccharides from each subunit displayed a distinct anion exchange HPLC profile due to a specific pattern of sialylation and sulfation. Six species were obtained from TSH alpha (with two glycosylation sites), including neutral oligosaccharides as well as those with one or two negative charges. For TSH beta (with one glycosylation site) at least eight oligosaccharide species were noted, representing nearly every permutation of sialylation and sulfation; approximately 30% contained three or more negative charges. Analysis of (3H)mannose-labeled oligosaccharides on Concanavalin-A-agarose showed 85% binding for those from TSH alpha, 70% for free alpha, and 50% for those from TSH beta. These data demonstrate that oligosaccharides from secreted TSH beta were more sialylated and sulfated, consistent with a more complex branching pattern, than those from TSH alpha. Oligosaccharides from free alpha-subunit were more sialylated than those from TSH alpha, and the net negative charge was intermediate between those of TSH alpha and TSH beta. Although great microheterogeneity is present even at the single glycosylation site on the beta-subunit of secreted TSH, a pattern of sialylation and sulfation could be discerned.
- Research Organization:
- National Institute of Diabetes and Digestive and Kidney Diseases, Bethesda, MD (USA)
- OSTI ID:
- 5879593
- Journal Information:
- Endocrinology; (United States), Vol. 124:6
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
OLIGOSACCHARIDES
CHEMICAL COMPOSITION
TSH
SECRETION
ASPARAGINE
CONCANAVALIN
DOUBLE LABELLING
HYPOTHYROIDISM
IN VITRO
ION EXCHANGE CHROMATOGRAPHY
LIQUID COLUMN CHROMATOGRAPHY
MANNOSE
MICE
PITUITARY GLAND
SULFATES
SULFUR 35
THYROID
TRACER TECHNIQUES
AGGLUTININS
ALDEHYDES
AMIDES
AMINO ACIDS
ANIMALS
ANTIBODIES
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BODY
CARBOHYDRATES
CARBOXYLIC ACIDS
CHROMATOGRAPHY
DAYS LIVING RADIOISOTOPES
DISEASES
ENDOCRINE DISEASES
ENDOCRINE GLANDS
EVEN-ODD NUCLEI
GLANDS
HEMAGGLUTININS
HEXOSES
HORMONES
ISOTOPE APPLICATIONS
ISOTOPES
LABELLING
LECTINS
LIGHT NUCLEI
MAMMALS
MONOSACCHARIDES
NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORGANS
OXYGEN COMPOUNDS
PEPTIDE HORMONES
PITUITARY HORMONES
RADIOISOTOPES
RODENTS
SACCHARIDES
SEPARATION PROCESSES
SULFUR COMPOUNDS
SULFUR ISOTOPES
VERTEBRATES
550201* - Biochemistry- Tracer Techniques