100% protein sequence coverage: a modern form of surrealism in proteomics

Amino Acids. 2011 Jul;41(2):291-310. doi: 10.1007/s00726-010-0680-6. Epub 2010 Jul 13.

Abstract

This review intends not only to discuss the current possibilities to gain 100% sequence coverage for proteins, but also to point out the critical limits to such an attempt. The aim of 100% sequence coverage, as the review title already implies, seems to be rather surreal if the complexity and dynamic range of a proteome is taken into consideration. Nevertheless, established bottom-up shotgun approaches are able to roughly identify a complete proteome as exemplary shown by yeast. However, this proceeding ignores more or less the fact that a protein is present as various protein species. The unambiguous identification of protein species requires 100% sequence coverage. Furthermore, the separation of the proteome must be performed on the protein species and not on the peptide level. Therefore, top-down is a good strategy for protein species analysis. Classical 2D-electrophoresis followed by an enzymatic or chemical cleavage, which is a combination of top-down and bottom-up, is another interesting approach. Moreover, the review summarizes further top-down and bottom-up combinations and to which extent middle-down improves the identification of protein species. The attention is also focused on cleavage strategies other than trypsin, as 100% sequence coverage in bottom-up experiments is only obtainable with a combination of cleavage reagents.

Publication types

  • Research Support, Non-U.S. Gov't
  • Review

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Humans
  • Mass Spectrometry / methods
  • Molecular Sequence Data
  • Peptide Fragments / chemistry
  • Protein Processing, Post-Translational*
  • Proteins / chemistry*
  • Proteomics / methods*
  • Sequence Analysis, Protein / methods*

Substances

  • Peptide Fragments
  • Proteins