|MadSci Network: Biochemistry|
There are two main approaches currently used in the characterization of proteins my mass spectrometry: top-down and bottom-up.
Top-down simply refers to direct mass spectrometric analysis of protein extracts or samples by virtue of the ability of the mass spectrometer to resolve the various forms of the proteins present in the sample. This can only be achieved with the use of high-resolution mass spectrometers.
Because intact proteins mixtures in a particular sample may contain many forms of the same proteins (for example, glycosylated and/or otherwise modified) and these forms may have very similar molecular weights, the mass spectrometer has to have the ability to distinguish these many forms. For this purpose only instruments such as TOF-TOF, FTICR (Fourier Transform ion cyclotron resonance) mass spectrometers or more recently Orbitraps can afford the resolution needed.
In bottom-up characterization, the samples are digested with enzymes to break down the proteins into amino acid clusters, while samples are not enzymatically digested in top-down characterization. Bottom-up characterization uses the amino acid information determined by mass spectrometry to build-up the proteins from which they came from.
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