Native top-down electrospray ionization-mass spectrometry of 158 kDa protein complex by high-resolution Fourier transform ion cyclotron resonance mass spectrometry.

Fourier transform ion cyclotron resonance mass spectrometry (FTICR MS) delivers high resolving power, mass measurement accuracy, and the capabilities for unambiguously sequencing by a top-down MS approach. Here, we report isotopic resolution of a 158 kDa protein complex, tetrameric aldolase with an average absolute deviation of 0.36 ppm and an average resolving power of ∼520 000 at m/z 6033 for the 26+ charge state in magnitude mode. Phase correction further improves the resolving power and average absolute deviation by 1.3-fold. Furthermore, native top-down electron capture dissociation (ECD) enables the sequencing of 168 C-terminal amino acid (AA) residues out of 463 total AAs. Combining the data from top-down MS of native and denatured aldolase complexes, a total of 56% of the total backbone bonds were cleaved. The observation of complementary product ion pairs confirms the correctness of the sequence and also the accuracy of the mass fitting of the isotopic distribution of the aldolase tetramer. Top-d...