Evaluation of bottlenecks in proinsulin secretion by Escherichia coli

• Tipo: Artigo em Revista Científica Internacional
• Data: 2004

Autores

Mergulhao, FJM (Autor) Outra; Taipa, MA (Autor) Outra; Cabral, JMS (Autor) Outra; Monteiro, GA (Autor) Outra

Revista

Título: Journal of Biotechnology

Vol. 109

Páginas: 31-43

ISSN: 0168-1656

Editora: Elsevier

Indexação

ISI Web of Knowledge - 23 Citações

ISI Web of Science

Scopus - 27 Citações

Classificação Científica

FOS: Ciências exactas e naturais > Ciências biológicas

CORDIS: Ciências Tecnológicas > Tecnologia > Biotecnologia

Outras Informações

ID Authenticus: P-000-AV1

DOI: 10.1016/j.jbiotec.2003.10.024

Abstract (EN): This work evaluates three potential bottlenecks in recombinant human proinsulin secretion by Escherichia coli: protein stability, secretion capacity and the effect of molecular size on secretion efficiency. A maximum secretion level of 7.2 mg g(-1) dry cell weight was obtained in the periplasm of E. coli JM109(DE3) host cells. This value probably represents an upper limit in the transport capacity of E. coli cells secreting ZZ-proinsulin and similar proteins with the protein A signal peptide. A selective deletion study was performed in the fusion partner and no effect of the molecular size (17-24 kDa) was detected on secretion efficiency. The protective effect against proteolysis provided by the ZZ domain was thoroughly demonstrated in the periplasm of E. coli and it was also shown that a single Z domain is able to provide the same protection level without compromising the downstream processing. The use of this shorter fusion partner enables a 1.6-fold increase in the recovery of the target protein after cleavage of the affinity handle.

Idioma: Inglês

Tipo (Avaliação Docente): Científica

Nº de páginas: 13