TY - JOUR
T1 - Heterologous expression and characterization of the aspartic endoprotease Pep4um from Ustilago maydis, a homolog of the human Chatepsin D, an important breast cancer therapeutic target
AU - Juárez-Montiel, Margarita
AU - Tesillo-Moreno, Pedro
AU - Cruz-Angeles, Ana
AU - Soberanes-Gutiérrez, Valentina
AU - Chávez-Camarillo, Griselda
AU - Ibarra, J. Antonio
AU - Hernández-Rodríguez, César
AU - Villa-Tanaca, Lourdes
N1 - Publisher Copyright:
© 2018, Springer Nature B.V.
PY - 2018/10/1
Y1 - 2018/10/1
N2 - The pep4um gene (um04926) of Ustilago maydis encodes a protein related to either vacuolar or lysosomal aspartic proteases. Bioinformatic analysis of the Pep4um protein revealed that it is a soluble protein with a signal peptide suggesting that it likely passes through the secretory pathway, and it has two probable self-activation sites, which are similar to those in Saccharomyces cerevisiae PrA. Moreover, the active site of the Pep4um has the two characteristic aspartic acid residues of aspartyl proteases. The pep4um gene was cloned, expressed in Pichia pastoris and a 54 kDa recombinant protein was observed. Pep4um-rec was confirmed to be an aspartic protease by specifically inhibiting its enzymatic activity with pepstatin A. Pep4um-rec enzymatic activity on acidic hemoglobin was optimal at pH 4.0 and at 40 °C. To the best of our knowledge this is the first report about the heterologous expression of an aspartic protease from a basidiomycete. An in-depth in silico analysis suggests that Pep4um is homolog of the human cathepsin D protein. Thus, the Pep4um-rec protein may be used to test inhibitors of human cathepsin D, an important breast cancer therapeutic target.
AB - The pep4um gene (um04926) of Ustilago maydis encodes a protein related to either vacuolar or lysosomal aspartic proteases. Bioinformatic analysis of the Pep4um protein revealed that it is a soluble protein with a signal peptide suggesting that it likely passes through the secretory pathway, and it has two probable self-activation sites, which are similar to those in Saccharomyces cerevisiae PrA. Moreover, the active site of the Pep4um has the two characteristic aspartic acid residues of aspartyl proteases. The pep4um gene was cloned, expressed in Pichia pastoris and a 54 kDa recombinant protein was observed. Pep4um-rec was confirmed to be an aspartic protease by specifically inhibiting its enzymatic activity with pepstatin A. Pep4um-rec enzymatic activity on acidic hemoglobin was optimal at pH 4.0 and at 40 °C. To the best of our knowledge this is the first report about the heterologous expression of an aspartic protease from a basidiomycete. An in-depth in silico analysis suggests that Pep4um is homolog of the human cathepsin D protein. Thus, the Pep4um-rec protein may be used to test inhibitors of human cathepsin D, an important breast cancer therapeutic target.
KW - Aspartyl acid endoprotease
KW - Cathepsin D homolog
KW - Pep4um recombinat protein
KW - Pepstatin A
KW - Pichia pastoris
KW - Ustilago maydis
UR - http://www.scopus.com/inward/record.url?scp=85051210119&partnerID=8YFLogxK
U2 - 10.1007/s11033-018-4267-8
DO - 10.1007/s11033-018-4267-8
M3 - Artículo
C2 - 30076522
AN - SCOPUS:85051210119
SN - 0301-4851
VL - 45
SP - 1155
EP - 1163
JO - Molecular Biology Reports
JF - Molecular Biology Reports
IS - 5
ER -