Issue 38, 2020

Fast protein analysis enabled by high-temperature hydrolysis


While the bottom-up protein analysis serves as a mainstream method for biological studies, its efficiency is limited by the time-consuming process for enzymatic digestion or hydrolysis as well as the post-digestion treatment prior to mass spectrometry analysis. In this work, we developed an enzyme-free microreaction system for fast and selective hydrolysis of proteins, and a direct analysis of the protein digests was achieved by nanoESI (electrospray ionization) mass spectrometry. Using the microreactor, proteins in aqueous solution could be selectively hydrolyzed at the aspartyl sites within 2 min at high temperatures (∼150 °C). Being free of salts, the protein digest solution could be directly analyzed using a mass spectrometer with nanoESI without further purification or post-digestion treatment. This method has been validated for the analysis of a variety of proteins with molecular weights ranging from 8.5 to 67 kDa. With introduction of a reducing agent into the protein solutions, fast cleavage of disulfide bonds was also achieved along with high-temperature hydrolysis, allowing for fast analysis of large proteins such as bovine serum albumin. The high-temperature microreaction system was also used with a miniature mass spectrometer for the determination of highly specific peptides from Mycobacterium tuberculosis antigens, showing its potential for point-of-care analysis of protein biomarkers.

Graphical abstract: Fast protein analysis enabled by high-temperature hydrolysis

Supplementary files

Article information

Article type
Edge Article
10 Jun 2020
09 Sep 2020
First published
10 Sep 2020
This article is Open Access

All publication charges for this article have been paid for by the Royal Society of Chemistry
Creative Commons BY license

Chem. Sci., 2020,11, 10506-10516

Fast protein analysis enabled by high-temperature hydrolysis

Y. Wang, W. Zhang and Z. Ouyang, Chem. Sci., 2020, 11, 10506 DOI: 10.1039/D0SC03237A

This article is licensed under a Creative Commons Attribution 3.0 Unported Licence. You can use material from this article in other publications without requesting further permissions from the RSC, provided that the correct acknowledgement is given.

Read more about how to correctly acknowledge RSC content.

Social activity