A 12-kilodalton N-glycosylated growth hormone-related peptide is present in human pituitary extracts.
This study was designed to investigate whether N-linked glycosylation could account for the presence of glycosylated GH forms (G-GH) in human pituitary extracts. The study was carried out in commercially available pituitary GH preparations (pitGH). Recombinant GHs obtained from eu- or prokaryotic cells were used as controls. Radioiodinated GHs were incubated in tubes containing Concanavalin-A (Con-A) attached to a Sepharose 4B matrix. Pituitary G-GH forms were selectively displaced from Con-A by adding N-acetyl-D-glucosamine or methyl-alpha-D-mannopyranoside and electrophoresed. Autoradiographies of these gels identified a 12-kilodalton (12K) band as the glycosylated form. Due to the fact that this peptide was partially immunoprecipitated with an anti-GH serum and the absence of detectable PRL in the pitGH extracts, it would indicate that such a glycopeptide was GH related. Endoglycosidase-F treatment of pitGH extracts induced a decrease in the mol wt of that 12K peptide, as indicated by the changes observed in its mobility on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This demonstrated that the sugar moieties were N-linked. When recombinant GHs were assayed by a similar method, no specific binding to Con-A was detected. NH2-terminal amino acid sequence analysis from the 12K band demonstrated that this band was composed by three peptides. Peptide 1 corresponds to the GH-N 102-119 sequence. Interestingly, peptide 2 exhibits GH-V 1-18 sequence, while peptide 3 seems to be a novel GH-related peptide. Taken together, these data suggest that the pituitary G-GH form found in human pituitary extracts is derived not from the "normal" GH-N gene, but, rather, from the GH-V gene or another unidentified gene. The potentially important pathophysiological implications of this finding need to be investigated.
Diaz
M J
MJ
Laboratorio de Neurociencias, Ramón Domínguez, Department of Physiology, Faculty of Medicine, Santiago de Compostela, Spain.
Dominguez
F
F
Haro
L S
LS
Ling
N
N
Devesa
J
J
D000595:Amino Acid Sequence; D003208:Concanavalin A; D004591:Electrophoresis, Polyacrylamide Gel; D006031:Glycosylation; D013006:Growth Hormone / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D007163:Immunosorbent Techniques; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase / Q000494:pharmacology; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010902:Pituitary Gland / Q000737:chemistry*
D003208:Concanavalin A; D013006:Growth Hormone; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase