Quantifying homologous proteins and proteoforms
This addresses the challenge of accurately measuring proteoforms for biologists studying protein functions, though it appears incremental as it builds on existing mass spectrometry methods.
The researchers tackled the problem of quantifying proteoform stoichiometries, which is hindered by peptide-specific biases in bottom-up mass spectrometry, and developed HIquant, a first-principles model that demonstrated high accuracy in quantifying fractional PTM occupancy without external standards, even for histone modifications.
Many proteoforms - arising from alternative splicing, post-translational modifications (PTMs), or paralogous genes - have distinct biological functions, such as histone PTM proteoforms. However, their quantification by existing bottom-up mass-spectrometry (MS) methods is undermined by peptide-specific biases. To avoid these biases, we developed and implemented a first-principles model (HIquant) for quantifying proteoform stoichiometries. We characterized when MS data allow inferring proteoform stoichiometries by HIquant, derived an algorithm for optimal inference, and demonstrated experimentally high accuracy in quantifying fractional PTM occupancy without using external standards, even in the challenging case of the histone modification code. HIquant server is implemented at: https://web.northeastern.edu/slavov/2014_HIquant/