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

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


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.
Author Shifeng Guo
Shifeng Guo,,
, Xiaoying Zhu
Xiaoying Zhu,,
, Dominik Jańczewski LPT
Dominik Jańczewski,,
- Laboratory Of Technological Processes
, Serina Siew Chen Lee
Serina Siew Chen Lee,,
, Tao He
Tao He,,
, Serena Lay Ming Teo
Serena Lay Ming Teo,,
, G. Julius Vancso
G. Julius Vancso,,
Journal seriesNature Nanotechnology, ISSN 1748-3387
Issue year2016
Publication size in sheets0.5
Keywords in PolishAFM, punkt izoelektryczny protein, LbL
Keywords in Englishnanobiotechnology; sensors
Languageen angielski
2016_NatureNano_Guo.pdf 3.63 MB
Score (nominal)50
ScoreMinisterial score = 50.0, 28-11-2017, ArticleFromJournal
Ministerial score (2013-2016) = 50.0, 28-11-2017, ArticleFromJournal
Publication indicators WoS Impact Factor: 2016 = 38.986 (2) - 2016=44.323 (5)
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