Decrease of hirudin degradation by deleting the KEX1 gene in recombinant Pichia pastoris
Research article published in Yeast (Chichester, England) (2008)
Abstract
Our previous study on recombinant hirudin production in Pichia pastoris demonstrated that, although the total productivity of hirudin was fairly high, its degradation was still severe, even if many engineering methods were applied to improve cell viability and reduce the release of intracellular proteinases. In this work, a pop-in/pop-out method, replacing the auxotrophic marker ARG4 gene with the resistant marker sh ble gene, was used to delete the KEX1 gene to reduce hirudin degradation in P. pastoris GS115Hir. Using this strategy, hirudin degradation was greatly decreased. At the same wet cell weight and cell viability, the percentage of intact hirudin Hir65 in total hirudin in strain GS115HirDeltakex1 was always kept as high as 90% in the initial stage of the methanol fermentation phase and above 62% even in the later stage of the methanol fermentation phase, whereas the percentage for the undeleted strain GS115Hir was only about 40% in the whole methanol fermentation phase. As a result, the intact hirudin Hir65 concentration could maximally reach 2.4 g/l in GS115HirDeltakex1 while it was only 1.1 g/l in GS115Hir.
Abstract sourced from PubMed (NCBI) for the cited record. See the original publication for the authoritative version.
Resumen
Our previous study on recombinant hirudin production in Pichia pastoris demonstrated that, although the total productivity of hirudin was fairly high, its degradation was still severe, even if many engineering methods were applied to improve cell viability and reduce the release of intracellular proteinases.
Por qué esto importa para la hirudoterapia
Este estudio examinó la producción de hirudina recombinante en Pichia pastoris, probando específicamente si la deleción del gen KEX1 podría reducir la degradación de la hirudina intacta (Hir65) durante la fermentación con metanol. La cepa con deleción de KEX1 mantuvo la hirudina intacta en aproximadamente el 90% inicialmente y por encima del 62% en etapas posteriores de la fermentación, frente a solo ~40% en la cepa sin deleción, alcanzando una concentración de hirudina intacta de 2,4 g/L en comparación con 1,1 g/L. Esto es altamente relevante para el ámbito de ASH porque aborda un cuello de botella biotecnológico práctico en la fabricación de un anticoagulante clave derivado de la sanguijuela. Sin embargo, se trata de un estudio de microbiología industrial y optimización de fermentación sin datos clínicos ni en animales; su relevancia se limita a la producción recombinante más que a la aplicación terapéutica de la hirudoterapia.
Citación
Decrease of hirudin degradation by deleting the KEX1 gene in recombinant Pichia pastoris
Ni Z et al. · Yeast (Chichester, England), 2008
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Añadido a la biblioteca ASH: May 27, 2026 · Última actualización del sitio: 18 de junio de 2026