Access

You are not currently logged in.

Access your personal account or get JSTOR access through your library or other institution:

login

Log in to your personal account or through your institution.

If You Use a Screen Reader

This content is available through Read Online (Free) program, which relies on page scans. Since scans are not currently available to screen readers, please contact JSTOR User Support for access. We'll provide a PDF copy for your screen reader.

Substrate Specificity of Barley Cysteine Endoproteases EP-A and EP-B

Anne Davy, Ib Svendsen, Susanne O. Sørensen, Mikael Blom Sørensen, Jacques Rouster, Morten Meldal, David J. Simpson and Verena Cameron-Mills
Plant Physiology
Vol. 117, No. 1 (May, 1998), pp. 255-261
Stable URL: http://www.jstor.org/stable/4278275
Page Count: 7
  • Read Online (Free)
  • Subscribe ($19.50)
  • Cite this Item
Since scans are not currently available to screen readers, please contact JSTOR User Support for access. We'll provide a PDF copy for your screen reader.
Substrate Specificity of Barley Cysteine Endoproteases EP-A and EP-B
Preview not available

Abstract

The cysteine endoproteases (EP)-A and EP-B were purified from green barley (Hordeum vulgare L.) malt, and their identity was confirmed by N-terminal amino acid sequencing. EP-B cleavage sites in recombinant type-C hordein were determined by N-terminal amino acid sequencing of the cleavage products, and were used to design internally quenched, fluorogenic peptide substrates. Tetrapeptide substrates of the general formula 2-aminobenzoyl-$\text{P}_{2}-\text{P}_{1}-\text{P}_{1}{}^{\prime}-\text{P}_{2}{}^{\prime}$-tyrosine(NO2)-aspartic acid, in which cleavage occurs between P1 and P1 ′, showed that the cysteine EPs preferred phenylalanine, leucine, or valine at P2. Arginine was preferred to glutamine at P1, whereas proline at P2, P1, or P1 ′ greatly reduced substrate kinetic specificity. Enzyme cleavage of C hordein was mainly determined by the primary sequence at the cleavage site, because elongation of substrates, based on the C hordein sequence, did not make them more suitable substrates. Site-directed mutagenesis of C hordein, in which serine or proline replaced leucine, destroyed primary cleavage sites. EP-A and EP-B were both more active than papain, mostly because of their much lower Km values.

Page Thumbnails

  • Thumbnail: Page 
255
    255
  • Thumbnail: Page 
256
    256
  • Thumbnail: Page 
257
    257
  • Thumbnail: Page 
258
    258
  • Thumbnail: Page 
259
    259
  • Thumbnail: Page 
260
    260
  • Thumbnail: Page 
261
    261