A strong nitrogen source-regulated promoter for controlled expression of foreign genes in the yeast Pichia pastoris

SG Shen, G Sulter, TW Jeffries, JM Cregg*

*Bijbehorende auteur voor dit werk

OnderzoeksoutputAcademicpeer review

116 Citaten (Scopus)

Samenvatting

In methylotrophic yeasts, glutathione-dependent formaldehyde dehydrogenase (FLD) is a key enzyme required for the metabolism of methanol as a carbon source and certain alkylated amines such as methylamine as nitrogen sources. We describe the isolation and characterization of the FLD1 gene from the yeast Pichia pastoris. The gene contains a single short intron with typical yeast-splicing signals near its 5' end, the first intron to be demonstrated in this yeast. The predicted FLD1 product (Fld1p) is a protein of 379 amino acids (approx. 40 kDa) with 71% identity to the FLD protein sequence from the n-alkane-assimilating yeast Candida maltosa and 61-65% identity with dehydrogenase class III enzymes from humans and other higher eukaryotes. Using p-lactamase as a reporter, we show that the FLD1 promoter (P-FLD1) is strongly and independently induced by either methanol as sole carbon source (with ammonium sulfate as nitrogen source) or methylamine as sole nitrogen source (with glucose as carbon source). Furthermore, with either methanol or methylamine induction, levels of p-lactamase produced under control of P-FLD1 are comparable to those obtained with the commonly used alcohol oxidase I gene promoter (P-AOX1). Thus, P-FLD1 is an attractive alternative to P-AOX1 for expression of foreign genes in P. pastoris, allowing the investigator a choice of carbon (methanol) or nitrogen source (methylamine) regulation with the same expression strain. (C) 1998 Elsevier Science B.V. All rights reserved.

Originele taal-2English
Pagina's (van-tot)93-102
Aantal pagina's10
TijdschriftGene
Volume216
Nummer van het tijdschrift1
StatusPublished - 17-aug-1998

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