Identification of mannose 6-phosphate in two asparagine-linked sugar chains of recombinant transforming growth factor-beta 1 precursor.
Recombinant transforming growth factor-beta 1 (TGF-beta 1) precursor produced and secreted by a clone of Chinese hamster ovary cells was found to be glycosylated and phosphorylated. Treatment of 32P-labeled precursor protein with N-glycanase indicated that phosphate was incorporated into asparagine-linked complex carbohydrate moieties. Fractionation of 32P-labeled glycopeptides followed by amino acid sequence analysis indicated that greater than 95% of the label was incorporated into two out of three glycosylation sites at Asn-82 and Asn-136 of the TGF-beta 1 precursor. Two-dimensional electrophoretic analysis of acid hydrolyzed precursor protein and precursor protein-derived glycopeptides indicated that 32P was incorporated as mannose 6-phosphate. Binding studies with the purified receptor for mannose 6-phosphate indicated that the TGF-beta 1 precursor could bind to this receptor and the binding was specifically inhibited with mannose 6-phosphate.
Purchio
A F
AF
Oncogen, Seattle, Washington 98121.
Cooper
J A
JA
Brunner
A M
AM
Lioubin
M N
MN
Gentry
L E
LE
Kovacina
K S
KS
Roth
R A
RA
Marquardt
H
H
D000595:Amino Acid Sequence; D000818:Animals; D001216:Asparagine; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D006600:Hexosephosphates / Q000032:analysis*; D008360:Mannosephosphates / Q000032:analysis* / Q000378:metabolism; D008969:Molecular Sequence Data; D011498:Protein Precursors; D011506:Proteins / Q000235:genetics / Q000378:metabolism; D017527:Receptor, IGF Type 2; D011994:Recombinant Proteins / Q000378:metabolism; D016212:Transforming Growth Factor beta; D015291:Transforming Growth Factors
D002352:Carrier Proteins; D006600:Hexosephosphates; D008360:Mannosephosphates; D011498:Protein Precursors; D011506:Proteins; D017527:Receptor, IGF Type 2; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; C027693:mannose-6-phosphate; D001216:Asparagine; D015291:Transforming Growth Factors