IGF1 inclusion bodies: A QbD based process approach for efficient USP as well as early DSP unit operations.
J Biotechnol
; 312: 23-34, 2020 Mar 20.
Article
em En
| MEDLINE
| ID: mdl-32114153
ABSTRACT
E. coli is an attractive host organism for strong recombinant protein expression. It expresses products either as soluble protein or as inclusion bodies (IB). IBs are insoluble, mostly inactive aggregates. However, recent progress enabled the efficient refolding back into their bioactive structure. Targeted IB production processes have been designed based on their characteristic features such as high yields along with purity and their simple separation. More profound process knowledge is needed to reveal interacting parameters required for quality by design grounded process development. This enables strategies for simplifying and accelerating upstream as well as downstream procedures. We present a workflow for gathering deeper process knowledge by a design of experiment approach for improved IGF1 IB formation in relation to impurity concentration. An IB expression maximum of 19.8 mgIGF1·gDCW-1 was found at pH 6.5, 37 °C and an IPTG induction of >45 µmol gDCW-1 for 12 h. Subsequently, three refolding buffers were tested together with a nonwoven anion exchange adsorber filter module. Knowledge-based buffer selection enabled high impurity log reduction values (LRVEndotoxin = 4.9; LRVDNA = 4.8, LRVHCP = 0.1-1) as well as chromatography column guarding potential by using those adsorptive matrices. Furthermore, adsorptive filtration followed by tangential flow filtration proved to be a promising alternative for product concentration.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Proteínas Recombinantes
/
Fator de Crescimento Insulin-Like I
/
Corpos de Inclusão
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Escherichia coli
Idioma:
En
Revista:
J Biotechnol
Assunto da revista:
BIOTECNOLOGIA
Ano de publicação:
2020
Tipo de documento:
Article
País de afiliação:
Áustria