Nejvíce citovaný článek - PubMed ID 23674150
Penicillin G acylase from Achromobacter sp. CCM 4824 : an efficient biocatalyst for syntheses of beta-lactam antibiotics under conditions employed in large-scale processes
Strain Pantoea agglomerans JM1 was isolated from the soil of a microbiome that had been exposed to polluting pharmaceuticals. The bacterium exhibited enzymatic activities useful for the biotransformation of beta-lactams. The genome of the strain was assembled and described, and the gene encoding valacyclovir-like hydrolase was identified.
- Publikační typ
- časopisecké články MeSH
This study deals with the potential of Pichia pastoris X-33 for the production of penicillin G acylase (PGAA) from Achromobacter sp. CCM 4824. Synthetic gene matching the codon usage of P. pastoris was designed for intracellular and secretion-based production strategies and cloned into vectors pPICZ and pPICZα under the control of AOX1 promoter. The simple method was developed to screen Pichia transformants with the intracellularly produced enzyme. The positive correlation between acylase production and pga gene dosage for both expression systems was demonstrated in small scale experiments. In fed-batch bioreactor cultures of X-33/PENS2, an extracellular expression system, total PGAA expressed from five copies reached 14,880 U/L of an active enzyme after 142 h; however, 60% of this amount retained in the cytosol. The maximum PGAA production of 31,000 U/L was achieved intracellularly from nine integrated gene copies of X-33/PINS2 after 90 h under methanol induction. The results indicate that in both expression systems the production level of PGAA is similar but there is a limitation in secretion efficiency.
- MeSH
- Achromobacter genetika metabolismus MeSH
- bioreaktory mikrobiologie MeSH
- exprese genu MeSH
- genetické vektory MeSH
- genová dávka MeSH
- klonování DNA MeSH
- kodon genetika MeSH
- penicilinamidasa genetika metabolismus MeSH
- Pichia genetika metabolismus MeSH
- promotorové oblasti (genetika) MeSH
- průmyslová mikrobiologie metody MeSH
- rekombinantní proteiny genetika metabolismus MeSH
- transformace genetická MeSH
- Publikační typ
- časopisecké články MeSH
- Názvy látek
- kodon MeSH
- penicilinamidasa MeSH
- rekombinantní proteiny MeSH
