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The substrate specificity of Metarhizium anisopliae and Bos taurus carboxypeptidases A: Insights into their use as tools for the removal of affinity tags

  1. Author:
    Austin, B. P.
    Tozser, J.
    Bagossi, P.
    Tropea, J. E.
    Waugh, D. S.

  2. Author Address

    [Austin, Brian P.; Tropea, Joseph E.; Waugh, David S.] NCI, Macromol Crystallog Lab, Ctr Canc Res, Frederick, MD 21702 USA. [Toezser, Jozsef; Bagossi, Peter] Debrecen Univ Med, Fac Med, Dept Biochem & Mol Biol, H-4012 Debrecen, Hungary.;Waugh, DS, NCI, Macromol Crystallog Lab, Ctr Canc Res, POB B, Frederick, MD 21702 USA.;waughd@mail.nih.gov
    1. Year: 2011
    2. Date: May
  2. Journal: Protein Expression and Purification
    1. 77
    2. 1
    3. Pages: 53-61
  4. Type of Article: Article
  5. ISSN: 1046-5928
  1. Abstract:

    Carboxypeptidases may serve as tools for removal of C-terminal affinity tags. In the present study, we describe the expression and purification of an A-type carboxypeptidase from the fungal pathogen Metarhizium anisopliae (MeCPA) that has been genetically engineered to facilitate the removal of polyhistidine tags from the C-termini of recombinant proteins. A complete, systematic analysis of the specificity of MeCPA in comparison with that of bovine carboxypeptidase A (BoCPA) was carried out. Our results indicate that the specificity of the two enzymes is similar but not identical. Histidine residues are removed more efficiently by MeCPA. The very inefficient digestion of peptides with C-terminal lysine or arginine residues, along with the complete inability of the enzyme to remove a C-terminal proline, suggests a strategy for designing C-terminal affinity tags that can be trimmed by MeCPA (or BoCPA) to produce a digestion product with a homogeneous endpoint. Published by Elsevier Inc.

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External Sources

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  2. DOI: 10.1016/j.pep.2010.11.005
  3. View record in Web of Science®
  4. WOS: 000288284200007

Library Notes

  1. Fiscal Year: FY2010-2011
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