Heterogeneity in porcine pituitary luteinizing hormone: amino acid and carbohydrate analysis.
Pituitary LH from porcine pituitary glands was purified by a buffered ethanol extraction procedure, ion exchange on DEAE- and carboxymethyl-cellulose, and molecular exclusion on Sephacryl S-200. Purity was assessed by amino acid composition, N-terminal sequence, and polyacrylamide gel electrophoresis. Subunits were isolated by countercurrent distribution and reverse phase HPLC. Four major forms of the alpha-subunit were detected: 1-96 (50%), 3-96 (23%), 4-96 (16%), and 7-96 (11%). [The original sequence report described only the 7-96 form, but we have detected the other forms in our studies of porcine FSH and in this and other species of LH.] Comparable N-terminal heterogeneity was not observed for the beta-subunit. Additional heterogeneity was observed for both subunits, attributable to heterogeneity in the N-linked oligosaccharide moieties. The isolated subunits were submitted to detailed compositional carbohydrate analysis, using pulsed amperometric detection of the HPLC-resolved sugar monomers after trifluoroacetic acid hydrolysis. Sialic acid and sulfate esters were estimated on separate hydrolyzates. The compositional data suggest that the two alpha-subunit N-linked moieties are hybrid complex biantennary structures with sulfated N-acetylgalactosamine (40-50%). Sixty to 70% of the alpha-subunit oligosaccharides are fucosylated. The beta-subunit of porcine LH has a single glycosylation site, which contains a mixture of biantennary oligosaccharide chains (80-90%) ending in N-acetylgalactosamine, half of which are sulfated. The balance (10-20%) are hybrid chains ending in sialylated galactose. The majority of the oligosaccharide on the beta-subunit is fucosylated.
van Dijk
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Department of Biochemistry and Molecular Biology, University of Texas M.D. Anderson Cancer Center, Houston 77030.
Ward
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D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis*; D000818:Animals; D002240:Carbohydrate Sequence; D002241:Carbohydrates / Q000032:analysis*; D007986:Luteinizing Hormone / Q000032:analysis* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D010902:Pituitary Gland / Q000737:chemistry*; D013552:Swine
D000596:Amino Acids; D002241:Carbohydrates; D007986:Luteinizing Hormone