TY - JOUR
T1 - Zymobacter palmae pyruvate decarboxylase production process development: Cloning in Escherichia coli, fed-batch culture and purification
AU - Alcover, Natàlia
AU - Carceller, Albert
AU - Álvaro, Gregorio
AU - Guillén, Marina
PY - 2019/7/1
Y1 - 2019/7/1
N2 - © 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim Pyruvate decarboxylase (PDC) is responsible for the decarboxylation of pyruvate, producing acetaldehyde and carbon dioxide and is of high interest for industrial applications. PDC is a very powerful tool in the enzymatic synthesis of chiral amines by combining it with transaminases when alanine is used as amine donor. However, one of the main drawback that hampers its use in biocatalysis is its production and the downstream processing on scale. In this paper, a production process of PDC from Zymobacter palmae has been developed. The enzyme has been cloned and overexpressed in Escherichia coli. It is presented, for the first time, the evaluation of the production of recombinant PDC in a bench-scale bioreactor, applying a substrate-limiting fed-batch strategy which led to a volumetric productivity and a final PDC specific activity of 6942 U L−1h−1 and 3677 U gDCW−1 (dry cell weight). Finally, PDC was purified in fast protein liquid chromatography equipment by ion exchange chromatography. The developed purification process resulted in 100% purification yield and a purification factor of 3.8.
AB - © 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim Pyruvate decarboxylase (PDC) is responsible for the decarboxylation of pyruvate, producing acetaldehyde and carbon dioxide and is of high interest for industrial applications. PDC is a very powerful tool in the enzymatic synthesis of chiral amines by combining it with transaminases when alanine is used as amine donor. However, one of the main drawback that hampers its use in biocatalysis is its production and the downstream processing on scale. In this paper, a production process of PDC from Zymobacter palmae has been developed. The enzyme has been cloned and overexpressed in Escherichia coli. It is presented, for the first time, the evaluation of the production of recombinant PDC in a bench-scale bioreactor, applying a substrate-limiting fed-batch strategy which led to a volumetric productivity and a final PDC specific activity of 6942 U L−1h−1 and 3677 U gDCW−1 (dry cell weight). Finally, PDC was purified in fast protein liquid chromatography equipment by ion exchange chromatography. The developed purification process resulted in 100% purification yield and a purification factor of 3.8.
KW - PDC purification
KW - Zymobacter palmae
KW - fed-batch culture
KW - high cell density cultures
KW - pyruvate decarboxylase (PDC)
UR - http://www.mendeley.com/research/zymobacter-palmae-pyruvate-decarboxylase-production-process-development-cloning-escherichia-coli-fed
U2 - 10.1002/elsc.201900010
DO - 10.1002/elsc.201900010
M3 - Article
C2 - 32625027
SN - 1618-0240
VL - 19
SP - 502
EP - 512
JO - Engineering in Life Sciences
JF - Engineering in Life Sciences
IS - 7
ER -