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Proteins pp 683-688 | Cite as

Kinetic Product Analysis of Aspartyl Proteinases Utilizing New Synthetic Substrates and Reversed Phase HPLC

  • Ben M. Dunn
  • Melba Jimenez
  • Jeff Weidner
  • Michael Pennington
  • Mark Carter
  • Benne Parten

Abstract

Kinetic characterization of the Aspartyl Proteinase family of enzymes has been hampered by the lack of convenient substrates. The enzymatic preference for cleavage between two hydrophobic residues causes most small synthetic substrates to be very insoluble in aqueous buffers. Since these enzymes do not cleave terminal residues readily, p-nitroanilides or p-nitrophenyl esters are not useful as chromophoric substrates and most previous assays required measurement of newly generated amino termini with ninhydrin or fluorescamine.

Keywords

Aspartic Proteinase Synthetic Substrate HPLC Grade Methanol Aspartyl Proteinase Phenyl Ester 
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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References

  1. 1.
    B. M. Dunn, B. Kammermann, and K. R. McCurry, The Synthesis, Purification, and Evaluation of a New Chromophoric Substrate for Pepsin and Other Aspartyl Proteases, Analyt. Biochem. 138: 68 (1984).Google Scholar
  2. 2.
    J. C. Powers, A. D. Harley, and D. V. Myers, Subsite Specificity of Porcine Pepsin, in: “Acid Proteinases: Structure, Function, and Biology”, J. Tang, ed., Plenum, New York (1977).Google Scholar
  3. 3.
    T. Hofmann and R. S. Hodges, A New Chromophoric Substrate for Penicillopepsin and Other Fungal Aspartic Proteinases, Biochem. J. 203: 603 (1982).PubMedGoogle Scholar
  4. 4.
    B. M. Dunn, B. Parten, M. Jimenez, C. E. Rolph, M. Valler, and J. Kay, Interaction of Aspartic Proteinases with a New Series of Synthetic Substrates and with Inhibitors Based on the Propart of Porcine Pepsinogen in: “Aspartic Proteinases and Their Inhibitors”, V. Kostka, ed., Walter de Gruyter, Berlin (1985).Google Scholar
  5. 5.
    M. J. Valler, J. Kay, and B. M. Dunn, Hydrolysis of a Series of Synthetic Chromophoric Substrates by Aspartic Proteinases, Trans. of the Biochem. Sac. 13: (in press) (1985).Google Scholar
  6. 6.
    P. Lindroth, and K. Mopper, High Performance Liquid Chromatographic Determination of Subpicomole Amounts of Amino Acids by Precolumn Fluorescence Derivitization with o-Phthaldialdehyde, Analyt. Chem., 51: 1667 (1979).CrossRefGoogle Scholar

Copyright information

© Plenum Press, New York 1987

Authors and Affiliations

  • Ben M. Dunn
    • 1
  • Melba Jimenez
    • 1
  • Jeff Weidner
    • 1
  • Michael Pennington
    • 1
  • Mark Carter
    • 1
  • Benne Parten
    • 1
  1. 1.Department of Biochemistry & Molecular BiologyUniversity of Florida College of MedicineGainesvilleUSA

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