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Angela J. Espejo Mojica

Angela Mosquera

Alexander Rodríguez-López

Dennis Díaz

Laura Beltrán

Francy Liliana Hernández

Carlos J. Alméciga Díaz

Luis A. Barrera

Abstract

β-hexosaminidases (Hex) are dimeric enzymes involved in the lysosomal degradation of glycolipids and glycans. They are formed by α- and/or β-subunits encoded by HEXA and HEXB genes, respectively. Mutations in these genes lead to Tay Sachs or Sandhoff diseases, which are neurodegenerative disorders caused by the accumulation of non-degraded glycolipids. Although tissue-derived Hex have been widely characterized, limited information is available for recombinant β-hexosaminidases. In this study, human lysosomal recombinant Hex (rhHex-A, rhHex-B, and rhHex-S) were produced in the methylotrophic yeast Pichia pastoris GS115. The highest specific enzyme activities were 13,124 for rhHexA; 12,779 for rhHex-B; and 14.606 U.mg-1 for rhHex-S. These results were 25- to 50-fold higher than those obtained from normal human leukocytes. Proteins were purified and characterized at different pH and temperature conditions. All proteins were stable at acidic pH, and at4 °C and 37 °C. At 45 °C rhHex-S was completely inactivated, while rhHex-A and rhHex-B showed high stability. This study demonstrates P. pastoris GS115 potential for polymeric lysosomal enzyme production, and describes the characterization of recombinant β-hexosaminidases produced within the same host.

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Keywords

β-N-acetylhexosaminidases, characterization, Pichia pastoris, recombinant hexosaminidases, Sandhoff disease, Tay Sachs disease.

References
How to Cite
Espejo Mojica, A., Mosquera, A., Rodríguez-López, A., Díaz, D., Beltrán, L., Hernández, F., Alméciga Díaz, C., & Barrera, L. (2016). Characterization of recombinant human lysosomal beta-hexosaminidases produced in the methylotrophic yeast Pichia pastoris. Universitas Scientiarum, 21(3), 195-217. https://doi.org/10.11144/Javeriana.SC21-3.corh
Section
Biotecnología /Biotechnology / Biotecnologia
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