The propeptide of prohormone convertase PC2 acts as a transferable aggregation and membrane-association signal

G Jan, N A Taylor, K T Scougall, K Docherty, K I Shennan

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

Prohormone convertase 2 (PC2) is a subtilisin-like protease involved in the intracellular processing of prohormones and proneuropeptides. Like its substrates, it is synthesised as a prepropeptide which undergoes proteolysis during transit through the regulated secretory pathway. Previous studies have shown that aggregation and membrane association of proPC2 occurs in a calcium-dependent and pH-dependent manner and that the pro-region of PC2 may be involved in this process. These events may be involved in the sorting of proteins to the regulated secretory pathway. To investigate this further, we made a chimeric protein containing both the signal peptide and pro-region of PC2 and the N-terminal part of alpha1-antitrypsin, called pro2alpha1. PC2, alpha1-antitrypsin and pro2alpha1 were compared with regard to their membrane association and aggregation properties using, respectively, sucrose gradient centrifugation after expression in Xenopus oocytes, and an in vitro aggregation assay. The chimeric protein, pro2alpha1, underwent low-pH-dependent aggregation and membrane association similar to wild-type PC2. Membrane association occurred at pH 5.5 in the absence of calcium and at pH 6.0 in the presence of 10 mM calcium but not at pH 6.5 or 7.0. alpha1-antitrypsin, as expected of a constitutively secreted protein, did not aggregate at low pH, nor associate with membranes. Pro2alpha1 thus exhibits the membrane association and aggregation properties of PC2, confirming the role of the pro-region in these processes. A series of deletions were performed within the 84-residue propeptide in order to define the sequences involved. Deletion of amino acids 52-77 reduced aggregation but large deletions in the pro-region had only a minimal effect on membrane association. These data suggest that several regions within the propeptide are important in these events.
Original languageEnglish
Pages (from-to)41-6
Number of pages6
JournalEuropean Journal of Biochemistry
Volume257
Issue number1
Publication statusPublished - 1 Oct 1998

Fingerprint

Proprotein Convertase 2
Proprotein Convertases
Agglomeration
Association reactions
Membranes
Secretory Pathway
Calcium
Proteins
Proteolysis
Subtilisin
Centrifugation
Protein Transport
Protein Sorting Signals
Xenopus
Sorting
Oocytes
Sucrose
Assays
Peptide Hydrolases
Amino Acids

Keywords

  • Animals
  • Base Sequence
  • Calcium
  • DNA Primers
  • Hydrogen-Ion Concentration
  • Membrane Proteins
  • Proprotein Convertase 2
  • Protein Sorting Signals
  • Recombinant Fusion Proteins
  • Sequence Deletion
  • Subtilisins
  • Xenopus

Cite this

The propeptide of prohormone convertase PC2 acts as a transferable aggregation and membrane-association signal. / Jan, G; Taylor, N A; Scougall, K T; Docherty, K; Shennan, K I.

In: European Journal of Biochemistry, Vol. 257, No. 1, 01.10.1998, p. 41-6.

Research output: Contribution to journalArticle

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abstract = "Prohormone convertase 2 (PC2) is a subtilisin-like protease involved in the intracellular processing of prohormones and proneuropeptides. Like its substrates, it is synthesised as a prepropeptide which undergoes proteolysis during transit through the regulated secretory pathway. Previous studies have shown that aggregation and membrane association of proPC2 occurs in a calcium-dependent and pH-dependent manner and that the pro-region of PC2 may be involved in this process. These events may be involved in the sorting of proteins to the regulated secretory pathway. To investigate this further, we made a chimeric protein containing both the signal peptide and pro-region of PC2 and the N-terminal part of alpha1-antitrypsin, called pro2alpha1. PC2, alpha1-antitrypsin and pro2alpha1 were compared with regard to their membrane association and aggregation properties using, respectively, sucrose gradient centrifugation after expression in Xenopus oocytes, and an in vitro aggregation assay. The chimeric protein, pro2alpha1, underwent low-pH-dependent aggregation and membrane association similar to wild-type PC2. Membrane association occurred at pH 5.5 in the absence of calcium and at pH 6.0 in the presence of 10 mM calcium but not at pH 6.5 or 7.0. alpha1-antitrypsin, as expected of a constitutively secreted protein, did not aggregate at low pH, nor associate with membranes. Pro2alpha1 thus exhibits the membrane association and aggregation properties of PC2, confirming the role of the pro-region in these processes. A series of deletions were performed within the 84-residue propeptide in order to define the sequences involved. Deletion of amino acids 52-77 reduced aggregation but large deletions in the pro-region had only a minimal effect on membrane association. These data suggest that several regions within the propeptide are important in these events.",
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T1 - The propeptide of prohormone convertase PC2 acts as a transferable aggregation and membrane-association signal

AU - Jan, G

AU - Taylor, N A

AU - Scougall, K T

AU - Docherty, K

AU - Shennan, K I

PY - 1998/10/1

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N2 - Prohormone convertase 2 (PC2) is a subtilisin-like protease involved in the intracellular processing of prohormones and proneuropeptides. Like its substrates, it is synthesised as a prepropeptide which undergoes proteolysis during transit through the regulated secretory pathway. Previous studies have shown that aggregation and membrane association of proPC2 occurs in a calcium-dependent and pH-dependent manner and that the pro-region of PC2 may be involved in this process. These events may be involved in the sorting of proteins to the regulated secretory pathway. To investigate this further, we made a chimeric protein containing both the signal peptide and pro-region of PC2 and the N-terminal part of alpha1-antitrypsin, called pro2alpha1. PC2, alpha1-antitrypsin and pro2alpha1 were compared with regard to their membrane association and aggregation properties using, respectively, sucrose gradient centrifugation after expression in Xenopus oocytes, and an in vitro aggregation assay. The chimeric protein, pro2alpha1, underwent low-pH-dependent aggregation and membrane association similar to wild-type PC2. Membrane association occurred at pH 5.5 in the absence of calcium and at pH 6.0 in the presence of 10 mM calcium but not at pH 6.5 or 7.0. alpha1-antitrypsin, as expected of a constitutively secreted protein, did not aggregate at low pH, nor associate with membranes. Pro2alpha1 thus exhibits the membrane association and aggregation properties of PC2, confirming the role of the pro-region in these processes. A series of deletions were performed within the 84-residue propeptide in order to define the sequences involved. Deletion of amino acids 52-77 reduced aggregation but large deletions in the pro-region had only a minimal effect on membrane association. These data suggest that several regions within the propeptide are important in these events.

AB - Prohormone convertase 2 (PC2) is a subtilisin-like protease involved in the intracellular processing of prohormones and proneuropeptides. Like its substrates, it is synthesised as a prepropeptide which undergoes proteolysis during transit through the regulated secretory pathway. Previous studies have shown that aggregation and membrane association of proPC2 occurs in a calcium-dependent and pH-dependent manner and that the pro-region of PC2 may be involved in this process. These events may be involved in the sorting of proteins to the regulated secretory pathway. To investigate this further, we made a chimeric protein containing both the signal peptide and pro-region of PC2 and the N-terminal part of alpha1-antitrypsin, called pro2alpha1. PC2, alpha1-antitrypsin and pro2alpha1 were compared with regard to their membrane association and aggregation properties using, respectively, sucrose gradient centrifugation after expression in Xenopus oocytes, and an in vitro aggregation assay. The chimeric protein, pro2alpha1, underwent low-pH-dependent aggregation and membrane association similar to wild-type PC2. Membrane association occurred at pH 5.5 in the absence of calcium and at pH 6.0 in the presence of 10 mM calcium but not at pH 6.5 or 7.0. alpha1-antitrypsin, as expected of a constitutively secreted protein, did not aggregate at low pH, nor associate with membranes. Pro2alpha1 thus exhibits the membrane association and aggregation properties of PC2, confirming the role of the pro-region in these processes. A series of deletions were performed within the 84-residue propeptide in order to define the sequences involved. Deletion of amino acids 52-77 reduced aggregation but large deletions in the pro-region had only a minimal effect on membrane association. These data suggest that several regions within the propeptide are important in these events.

KW - Animals

KW - Base Sequence

KW - Calcium

KW - DNA Primers

KW - Hydrogen-Ion Concentration

KW - Membrane Proteins

KW - Proprotein Convertase 2

KW - Protein Sorting Signals

KW - Recombinant Fusion Proteins

KW - Sequence Deletion

KW - Subtilisins

KW - Xenopus

M3 - Article

C2 - 9799100

VL - 257

SP - 41

EP - 46

JO - European Journal of Biochemistry

JF - European Journal of Biochemistry

SN - 0014-2956

IS - 1

ER -