Direct Identification of Functional Amyloid Proteins by Label-Free Quantitative Mass Spectrometry
AbstractFunctional amyloids are important structural and functional components of many biofilms, yet our knowledge of these fascinating polymers is limited to a few examples for which the native amyloids have been isolated in pure form. Isolation of the functional amyloids from other cell components represents a major bottleneck in the search for new functional amyloid systems. Here we present a label-free quantitative mass spectrometry method that allows identification of amyloid proteins directly in cell lysates. The method takes advantage of the extreme structural stability and polymeric nature of functional amyloids and the ability of concentrated formic acid to depolymerize the amyloids. An automated data processing pipeline that provides a short list of amyloid protein candidates was developed based on an amyloid-specific sigmoidal abundance signature in samples treated with increasing concentrations of formic acid. The method was evaluated using the Escherichia coli curli and the Pseudomonas Fap system. It confidently identified the major amyloid subunit for both systems, as well as the minor subunit for the curli system. A few non-amyloid proteins also displayed the sigmoidal abundance signature. However, only one of these contained a sec-dependent signal peptide, which characterizes most of all secreted proteins, including all currently known functional bacterial amyloids. View Full-Text
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Danielsen, H.N.; Hansen, S.H.; Herbst, F.-A.; Kjeldal, H.; Stensballe, A.; Nielsen, P.H.; Dueholm, M.S. Direct Identification of Functional Amyloid Proteins by Label-Free Quantitative Mass Spectrometry. Biomolecules 2017, 7, 58.
Danielsen HN, Hansen SH, Herbst F-A, Kjeldal H, Stensballe A, Nielsen PH, Dueholm MS. Direct Identification of Functional Amyloid Proteins by Label-Free Quantitative Mass Spectrometry. Biomolecules. 2017; 7(3):58.Chicago/Turabian Style
Danielsen, Heidi N.; Hansen, Susan H.; Herbst, Florian-Alexander; Kjeldal, Henrik; Stensballe, Allan; Nielsen, Per H.; Dueholm, Morten S. 2017. "Direct Identification of Functional Amyloid Proteins by Label-Free Quantitative Mass Spectrometry." Biomolecules 7, no. 3: 58.