Atomic force microscope study of the effect of the immobilization substrate on the structure and force-extension curves of a multimeric protein

Jeffrey G. Forbes, Albert J. Jin, Kuan Wang

Research output: Contribution to journalArticlepeer-review

26 Citations (Scopus)

Abstract

Atomic force microscopy (AFM) of soluble proteins requires that the protein be immobilized on a flat substrate. Many different substrates have been used successfully for imaging proteins, with mica and gold on mica being the most commonly used. Successful imaging of a soluble protein requires that the protein adhere to the substrate; substrate adhesion is often the primary determinant in choosing a surface for AFM imaging. As a result of the constraint on substrate adhesion, little data is available on how the substrate can affect the conformation of proteins and the resulting images. The AdhE protein of Escherichia coli is a multienzyme that forms supramolecular structures composed of 20-60 subunits. We have used atomic force microscopy to study the aggregation state of the purified protein on different substrates and its elastic properties as measured by AFM. We have obtained both contact mode and noncontact (MAC mode) images of the protein assemblies immobilized on a substrate and imaged under buffer. Noncontact mode images on a mica surface show elongated structures as previously observed via electron microscopy, whereas the contact mode images on a gold surface over mica are dominated by globular particles composed of 1-8 monomers. Forced extension of the polypeptide chains yields force versus distance curves which may be fit with the wormlike chain (WLC) model. Our results are consistent with the subnanometer persistence length expected for a typical polypeptide chain. These results indicate that the extension of a protein chain in a supramolecular assembly is not significantly affected by the neighboring proteins.

Original languageEnglish
Pages (from-to)3067-3075
Number of pages9
JournalLangmuir
Volume17
Issue number10
DOIs
Publication statusPublished - May 15 2001
Externally publishedYes

ASJC Scopus subject areas

  • Materials Science(all)
  • Condensed Matter Physics
  • Surfaces and Interfaces
  • Spectroscopy
  • Electrochemistry

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