Structure of recombinant N-terminal globule of type VI collagen α3 chain and its binding to heparin and hyaluronan

Ulrich Specks, Ulrike Mayer, Roswitha Nischt, Thomas Spissinger, Karlheinz Mann, Rupert Timpl, Jürgen Engel, Mon Li Chu

Research output: Contribution to journalArticle

114 Citations (Scopus)

Abstract

A large portion of the N-terminal globule of human collagen VI was prepared from the culture medium of stably transfected human embryonic kidney cell clones. The recombinant product corresponds to sequence positions 1-1586 of the α3(VI) chain that consists of eight homologous ∼200 residue motifs (N9 to N2) being similar to the A domain motif of von Willebrand factor. By ultracentrifugation fragment N9-N2 showed a molecular mass of 180 kDa and an asymmetric shape. Elongated structures that consist of eight small globes (diameter ∼5 nm) were demonstrated by electron microscopy. The data indicate that each A domain motif represents a separate folding unit which are connected to each other by short protease-sensitive peptide segments. Circular dichroism studies demonstrated about 38% α helix, 14% β sheets and 17% β turns. Fragment N9-N2 showed binding to heparin which could be abolished by moderate salt concentrations. Heparin binding was assigned to domains N9, N6 and N3 which were obtained after partial proteolysis. Domains N7, N5 and N4 lacked affinity for heparin. In addition, N9-N2 showed strong binding to hyaluronan that required exposure to 6 M urea for full dissociation. Ligand binding studies indicated some affinity of N9-N2 for the triple helical region of collagen VI suggesting a role of the N-terminal globule in the self-assembly of microfibrils. No or only little binding was, however, observed to fibril-forming collagens I and III, several basement membrane proteins and other extracellular proteins. Fragment N9-N2 was also an inactive substrate for cell adhesion.

Original languageEnglish (US)
Pages (from-to)4281-4290
Number of pages10
JournalEMBO Journal
Volume11
Issue number12
StatePublished - 1992
Externally publishedYes

Fingerprint

Collagen Type VI
Collagen Type III
Hyaluronic Acid
Heparin
Collagen
Clone cells
Proteolysis
Microfibrils
Ultracentrifugation
Cell adhesion
von Willebrand Factor
Molecular mass
Circular Dichroism
Basement Membrane
Cell Adhesion
Self assembly
Electron microscopy
Culture Media
Urea
Electron Microscopy

Keywords

  • A domain motif
  • Collagen microfibrils
  • Eukaryotic expression vector
  • Glycosaminoglycans
  • Protein-protein and cell-matrix interactions

ASJC Scopus subject areas

  • Genetics
  • Cell Biology

Cite this

Specks, U., Mayer, U., Nischt, R., Spissinger, T., Mann, K., Timpl, R., ... Chu, M. L. (1992). Structure of recombinant N-terminal globule of type VI collagen α3 chain and its binding to heparin and hyaluronan. EMBO Journal, 11(12), 4281-4290.

Structure of recombinant N-terminal globule of type VI collagen α3 chain and its binding to heparin and hyaluronan. / Specks, Ulrich; Mayer, Ulrike; Nischt, Roswitha; Spissinger, Thomas; Mann, Karlheinz; Timpl, Rupert; Engel, Jürgen; Chu, Mon Li.

In: EMBO Journal, Vol. 11, No. 12, 1992, p. 4281-4290.

Research output: Contribution to journalArticle

Specks, U, Mayer, U, Nischt, R, Spissinger, T, Mann, K, Timpl, R, Engel, J & Chu, ML 1992, 'Structure of recombinant N-terminal globule of type VI collagen α3 chain and its binding to heparin and hyaluronan', EMBO Journal, vol. 11, no. 12, pp. 4281-4290.
Specks, Ulrich ; Mayer, Ulrike ; Nischt, Roswitha ; Spissinger, Thomas ; Mann, Karlheinz ; Timpl, Rupert ; Engel, Jürgen ; Chu, Mon Li. / Structure of recombinant N-terminal globule of type VI collagen α3 chain and its binding to heparin and hyaluronan. In: EMBO Journal. 1992 ; Vol. 11, No. 12. pp. 4281-4290.
@article{f675e8f98ed644ac85c30df23af8772d,
title = "Structure of recombinant N-terminal globule of type VI collagen α3 chain and its binding to heparin and hyaluronan",
abstract = "A large portion of the N-terminal globule of human collagen VI was prepared from the culture medium of stably transfected human embryonic kidney cell clones. The recombinant product corresponds to sequence positions 1-1586 of the α3(VI) chain that consists of eight homologous ∼200 residue motifs (N9 to N2) being similar to the A domain motif of von Willebrand factor. By ultracentrifugation fragment N9-N2 showed a molecular mass of 180 kDa and an asymmetric shape. Elongated structures that consist of eight small globes (diameter ∼5 nm) were demonstrated by electron microscopy. The data indicate that each A domain motif represents a separate folding unit which are connected to each other by short protease-sensitive peptide segments. Circular dichroism studies demonstrated about 38{\%} α helix, 14{\%} β sheets and 17{\%} β turns. Fragment N9-N2 showed binding to heparin which could be abolished by moderate salt concentrations. Heparin binding was assigned to domains N9, N6 and N3 which were obtained after partial proteolysis. Domains N7, N5 and N4 lacked affinity for heparin. In addition, N9-N2 showed strong binding to hyaluronan that required exposure to 6 M urea for full dissociation. Ligand binding studies indicated some affinity of N9-N2 for the triple helical region of collagen VI suggesting a role of the N-terminal globule in the self-assembly of microfibrils. No or only little binding was, however, observed to fibril-forming collagens I and III, several basement membrane proteins and other extracellular proteins. Fragment N9-N2 was also an inactive substrate for cell adhesion.",
keywords = "A domain motif, Collagen microfibrils, Eukaryotic expression vector, Glycosaminoglycans, Protein-protein and cell-matrix interactions",
author = "Ulrich Specks and Ulrike Mayer and Roswitha Nischt and Thomas Spissinger and Karlheinz Mann and Rupert Timpl and J{\"u}rgen Engel and Chu, {Mon Li}",
year = "1992",
language = "English (US)",
volume = "11",
pages = "4281--4290",
journal = "EMBO Journal",
issn = "0261-4189",
publisher = "Nature Publishing Group",
number = "12",

}

TY - JOUR

T1 - Structure of recombinant N-terminal globule of type VI collagen α3 chain and its binding to heparin and hyaluronan

AU - Specks, Ulrich

AU - Mayer, Ulrike

AU - Nischt, Roswitha

AU - Spissinger, Thomas

AU - Mann, Karlheinz

AU - Timpl, Rupert

AU - Engel, Jürgen

AU - Chu, Mon Li

PY - 1992

Y1 - 1992

N2 - A large portion of the N-terminal globule of human collagen VI was prepared from the culture medium of stably transfected human embryonic kidney cell clones. The recombinant product corresponds to sequence positions 1-1586 of the α3(VI) chain that consists of eight homologous ∼200 residue motifs (N9 to N2) being similar to the A domain motif of von Willebrand factor. By ultracentrifugation fragment N9-N2 showed a molecular mass of 180 kDa and an asymmetric shape. Elongated structures that consist of eight small globes (diameter ∼5 nm) were demonstrated by electron microscopy. The data indicate that each A domain motif represents a separate folding unit which are connected to each other by short protease-sensitive peptide segments. Circular dichroism studies demonstrated about 38% α helix, 14% β sheets and 17% β turns. Fragment N9-N2 showed binding to heparin which could be abolished by moderate salt concentrations. Heparin binding was assigned to domains N9, N6 and N3 which were obtained after partial proteolysis. Domains N7, N5 and N4 lacked affinity for heparin. In addition, N9-N2 showed strong binding to hyaluronan that required exposure to 6 M urea for full dissociation. Ligand binding studies indicated some affinity of N9-N2 for the triple helical region of collagen VI suggesting a role of the N-terminal globule in the self-assembly of microfibrils. No or only little binding was, however, observed to fibril-forming collagens I and III, several basement membrane proteins and other extracellular proteins. Fragment N9-N2 was also an inactive substrate for cell adhesion.

AB - A large portion of the N-terminal globule of human collagen VI was prepared from the culture medium of stably transfected human embryonic kidney cell clones. The recombinant product corresponds to sequence positions 1-1586 of the α3(VI) chain that consists of eight homologous ∼200 residue motifs (N9 to N2) being similar to the A domain motif of von Willebrand factor. By ultracentrifugation fragment N9-N2 showed a molecular mass of 180 kDa and an asymmetric shape. Elongated structures that consist of eight small globes (diameter ∼5 nm) were demonstrated by electron microscopy. The data indicate that each A domain motif represents a separate folding unit which are connected to each other by short protease-sensitive peptide segments. Circular dichroism studies demonstrated about 38% α helix, 14% β sheets and 17% β turns. Fragment N9-N2 showed binding to heparin which could be abolished by moderate salt concentrations. Heparin binding was assigned to domains N9, N6 and N3 which were obtained after partial proteolysis. Domains N7, N5 and N4 lacked affinity for heparin. In addition, N9-N2 showed strong binding to hyaluronan that required exposure to 6 M urea for full dissociation. Ligand binding studies indicated some affinity of N9-N2 for the triple helical region of collagen VI suggesting a role of the N-terminal globule in the self-assembly of microfibrils. No or only little binding was, however, observed to fibril-forming collagens I and III, several basement membrane proteins and other extracellular proteins. Fragment N9-N2 was also an inactive substrate for cell adhesion.

KW - A domain motif

KW - Collagen microfibrils

KW - Eukaryotic expression vector

KW - Glycosaminoglycans

KW - Protein-protein and cell-matrix interactions

UR - http://www.scopus.com/inward/record.url?scp=0026484847&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0026484847&partnerID=8YFLogxK

M3 - Article

C2 - 1425570

AN - SCOPUS:0026484847

VL - 11

SP - 4281

EP - 4290

JO - EMBO Journal

JF - EMBO Journal

SN - 0261-4189

IS - 12

ER -