Measuring protein isoelectric points by AFM-based force spectroscopy using trace amounts of sample

Shifeng Guo, Xiaoying Zhu, Dominik Jańczewski (Corresponding Author), Serina Siew Chen Lee, Tao He, Serena Lay Ming Teo, G. Julius Vancso (Corresponding Author)

Research output: Contribution to journalArticleAcademicpeer-review

21 Citations (Scopus)
1 Downloads (Pure)

Abstract

Protein charge at various pH and isoelectric point (pI) values is important in understanding protein function. However, often only trace amounts of unknown proteins are available and pI measurements cannot be obtained using conventional methods. Here, we show a method based on the atomic force microscope (AFM) to determine pI using minute quantities of proteins. The protein of interest is immobilized on AFM colloidal probes and the adhesion force of the protein is measured against a positively and a negatively charged substrate made by layer-by-layer deposition of polyelectrolytes. From the AFM force–distance curves, pI values with an estimated accuracy of ±0.25 were obtained for bovine serum albumin, myoglobin, fibrinogen and ribonuclease A over a range of 4.7–9.8. Using this method, we show that the pI of the ‘footprint’ of the temporary adhesive proteins secreted by the barnacle cyprid larvae of Amphibalanus amphitrite is in the range 9.6–9.7.
Original languageEnglish
Pages (from-to)817-823
JournalNature nanotechnology
Volume11
DOIs
Publication statusPublished - 2016

Keywords

  • METIS-318036
  • IR-101770

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