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Proteins pp 343-353 | Cite as

Amidation of Protein Carboxyl Groups

  • George E. Tarr

Abstract

One of the delights of working with proteins, as opposed to nucleic acids, is their great variation in properties. But for the protein sequencer concerned only with covalent structure, this is also a problem. For example, a protein which is acidic or easily precipitated by organic solvents may not elute from a reversed-phase HPLC column under the standard conditions of dilute acid and acetonitrile (MeCN) so effective in a majority of cases. Use of a neutral buffer (a UV-transparent, volatile one is available)1 alleviates problems due to acidity but aggrevates those due to precipitation. And many proteins that do behave well on HPLC may balk when treated with a protease: they are either insoluble in suitable buffers or present a refractory surface with susceptible cleavage sites buried. The usual stratagem is to include a denaturant, but the level of urea that trypsin and most other enzymes will tolerate is often insufficient to promote complete and repoducible digestion, and detergents — which also are not always effective — interfer with separations employing RP-HPLC. An alternative to adjusting conditions to try to accomodate each ill-behaved protein is to modify these proteins at the start in such a way that they are all alike in their properties and common problems are overcome. Ideally this modification should be specific, complete, stable, universally produced under gentle conditions, unrestrictive with regard to other manipulations, and advantageous during sequencing.

Keywords

Cyanogen Bromide Refractory Surface Insulin Chain Attachment Procedure Glycine Amide 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Copyright information

© Plenum Press, New York 1987

Authors and Affiliations

  • George E. Tarr
    • 1
  1. 1.UM Protein Sequencing Facility, Department of Biological ChemistryUniversity of MichiganAnn ArborUSA

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