Scanning Transmission Electron Microscopy and Small-Angle Scattering Provide Evidence That Native Escherichia coli ClpP Is a Tetradecamer with an Axial Pore

John Flanagan, Joseph S. Wall, Malcolm S. Capel, Dieter K. Schneider, John Shanklin

Research output: Contribution to journalArticle

43 Citations (Scopus)

Abstract

The Escherichia coli ATP-dependent caseinolytic protease (Clp) is composed of two distinct subunits; protease, ClpP, and ATPase, ClpA. Active ClpP has been overexpressed to approximately 50% of soluble protein in E. coli, and purified to homogeneity. Direct mass determination of individual particles using scanning transmission electron microscopy (STEM) yields a mean native molecular mass of 305 ± 9 kDa for the ClpP oligomer, suggesting that it has a tetradecameric structure. Small-angle X-ray scattering (SAXS) curves were determined for ClpP in solution at concentrations of 1 - 10 mg/mL. A combination of STEM and SAXS data was used to derive a model for ClpP, comprising a cylindrical oligomer about 100 Å in diameter and about 75 Å in height with an axial pore about 32-36 Å in diameter. The volume of the pore is estimated to be ~70 000 Å 3 , similar in size to those found in chaperone proteins, and is large enough to accommodate unfolded polypeptide chains, although most globular folded proteins would be excluded.

Original languageEnglish (US)
Pages (from-to)10910-10917
Number of pages8
JournalBiochemistry
Volume34
Issue number34
DOIs
StatePublished - Jan 1 1995

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Small Angle Scattering
Scanning Transmission Electron Microscopy
Escherichia coli
ATP-Dependent Proteases
X-Rays
Scattering
Transmission electron microscopy
X ray scattering
Oligomers
Scanning electron microscopy
Escherichia coli Proteins
Peptide Hydrolases
Adenosine Triphosphatases
Proteins
Molecular mass
Peptides
Adenosine Triphosphate

All Science Journal Classification (ASJC) codes

  • Biochemistry

Cite this

Flanagan, John ; Wall, Joseph S. ; Capel, Malcolm S. ; Schneider, Dieter K. ; Shanklin, John. / Scanning Transmission Electron Microscopy and Small-Angle Scattering Provide Evidence That Native Escherichia coli ClpP Is a Tetradecamer with an Axial Pore. In: Biochemistry. 1995 ; Vol. 34, No. 34. pp. 10910-10917.
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abstract = "The Escherichia coli ATP-dependent caseinolytic protease (Clp) is composed of two distinct subunits; protease, ClpP, and ATPase, ClpA. Active ClpP has been overexpressed to approximately 50{\%} of soluble protein in E. coli, and purified to homogeneity. Direct mass determination of individual particles using scanning transmission electron microscopy (STEM) yields a mean native molecular mass of 305 ± 9 kDa for the ClpP oligomer, suggesting that it has a tetradecameric structure. Small-angle X-ray scattering (SAXS) curves were determined for ClpP in solution at concentrations of 1 - 10 mg/mL. A combination of STEM and SAXS data was used to derive a model for ClpP, comprising a cylindrical oligomer about 100 {\AA} in diameter and about 75 {\AA} in height with an axial pore about 32-36 {\AA} in diameter. The volume of the pore is estimated to be ~70 000 {\AA} 3 , similar in size to those found in chaperone proteins, and is large enough to accommodate unfolded polypeptide chains, although most globular folded proteins would be excluded.",
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Scanning Transmission Electron Microscopy and Small-Angle Scattering Provide Evidence That Native Escherichia coli ClpP Is a Tetradecamer with an Axial Pore. / Flanagan, John; Wall, Joseph S.; Capel, Malcolm S.; Schneider, Dieter K.; Shanklin, John.

In: Biochemistry, Vol. 34, No. 34, 01.01.1995, p. 10910-10917.

Research output: Contribution to journalArticle

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