Biosynthesis and packaging of carboxypeptidase D into nascent secretory vesicles in pituitary cell lines

Oleg Varlamov, Fang Wu, Dennis Shields, Lloyd D. Fricker

Research output: Contribution to journalArticle

29 Citations (Scopus)

Abstract

Metallocarboxypeptidase D (CPD) is a membrane-bound trans. Golgi network (TGN) protein. In AtT-20 cells, CPD is initially produced as a 170-kDa endoglycosidase H-sensitive glycoprotein. Within 30 min of chase, the CPD increases to 180 kDa and is resistant to endoglycosidase H as a result of carbohydrate maturation. CPD also undergoes an activation step required for binding to a substrate affinity resin. Blocking the protein exit from the endoplasmic reticulum inhibits the increase in molecular mass but not the step required for affinity column binding, suggesting that enzyme activation precedes carbohydrate maturation and that these reactions occur in distinct intracellular compartments. Only the higher molecular weight mature CPD enters nascent secretory vesicles, which bud from the TGN of permeabilized AtT-20 and GH3 cells. The budding efficiency of CPD into vesicles is 2-3- fold lower than that of endogenous proopiomelanocortin in ART-20 cells or prolactin in GH3 cells. In contrast, the packaging of a truncated form of CPD, which lacks the cytoplasmic tall and transmembrane domain, was similar to that of proopiomelanocortin. Taken together, the results support the proposal that CPD functions in the TGN in the processing of proteins that transit the secretory pathway and that the C-terminal region plays a major role in TGN retention.

Original languageEnglish (US)
Pages (from-to)14040-14045
Number of pages6
JournalJournal of Biological Chemistry
Volume274
Issue number20
DOIs
StatePublished - May 14 1999
Externally publishedYes

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Biosynthesis
Secretory Vesicles
Product Packaging
Pro-Opiomelanocortin
Packaging
Glycoside Hydrolases
Cells
Cell Line
Chemical activation
Carbohydrates
Proteins
Molecular mass
Prolactin
Enzyme Activation
Secretory Pathway
Glycoproteins
Resins
Molecular weight
Endoplasmic Reticulum
Membranes

ASJC Scopus subject areas

  • Biochemistry

Cite this

Biosynthesis and packaging of carboxypeptidase D into nascent secretory vesicles in pituitary cell lines. / Varlamov, Oleg; Wu, Fang; Shields, Dennis; Fricker, Lloyd D.

In: Journal of Biological Chemistry, Vol. 274, No. 20, 14.05.1999, p. 14040-14045.

Research output: Contribution to journalArticle

Varlamov, Oleg ; Wu, Fang ; Shields, Dennis ; Fricker, Lloyd D. / Biosynthesis and packaging of carboxypeptidase D into nascent secretory vesicles in pituitary cell lines. In: Journal of Biological Chemistry. 1999 ; Vol. 274, No. 20. pp. 14040-14045.
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