The ICL1 gene of Pichia pastoris, transcriptional regulation and use of its promoter.
(2003)
Journal - Yeast (Chichester, England) (England )
Abstract :
We cloned and characterized a gene encoding isocitrate lyase from the methylotrophic yeast Pichia pastoris. This gene was isolated from a P. pastoris genomic library using a homologous PCR hybridization probe, amplified with two sets of degenerate primers designed from conserved regions in yeast isocitrate lyases. The cloned gene was sequenced and consists of an open reading frame of 1563 bp encoding a protein of 551 amino acids. The molecular mass of the protein is calculated to be 60.6 kDa with high sequence similarity to isocitrate lyase from other organisms. There is a 64% identity between amino acid sequences of P. pastoris Icl and Saccharomyces cerevisiae Icl. Northern blot analyses showed that, as in S. cerevisiae, the steady-state ICL1 mRNA levels depend on the carbon source used for cell growth. Expression in P. pastoris of the dextranase gene (dexA) from Penicillium minioluteum under control of the ICL1 promoter proved that P(ICL1) is a good alternative for the expression of heterologous proteins in this methylotrophic yeast. The sequence presented here has been deposited in the EMBL data library under Accession No. AJ272040.Copyright 2003 John Wiley & Sons, Ltd.
| ISSN : | 0749-503X |
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| Mesh Heading : | Amino Acid Sequence Base Sequence Blotting, Northern Cloning, Molecular Dextranase Fungal Proteins Gene Expression Regulation, Fungal Isocitrate Lyase Molecular Sequence Data Molecular Weight Mutagenesis, Insertional Pichia Promoter Regions, Genetic Sequence Alignment Transcription, Genetic genetics metabolism metabolism metabolism metabolism physiology |
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| Mesh Heading Relevant : | genetics genetics genetics genetics physiology |
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Isolation of the Pichia pastoris PYC1 gene encoding pyruvate carboxylase and identification of a suppressor of the pyc phenotype.
(1998)
Journal - Yeast (Chichester, England) (ENGLAND )
Abstract :
We have cloned and characterized a gene encoding pyruvate carboxylase from the methylotrophic yeast Pichia pastoris. Disruption of this gene produced inability to grow in minimal medium with glucose as carbon source and ammonium as nitrogen source. Growth was possible with aspartate or glutamate as nitrogen source. The gene PpPYC1 expressed from its own promoter was able to rescue the phenotype of Saccharomyces cerevisiae mutants devoid of pyruvate carboxylase. In a P. pastoris strain carrying a disrupted PpPYC1 gene we have isolated spontaneous mutants able to grow in non-permissive conditions. In a mutant strain grown in glucose several enzymes sensitive to catabolite repression were derepressed. The strain also had elevated levels of glutamate dehydrogenase (NAD) both in repressed and derepressed conditions.
| ISSN : | 0749-503X |
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| Mesh Heading : | Amino Acid Sequence Base Sequence Culture Media Gene Expression Regulation, Fungal Molecular Sequence Data Phenotype Pichia Pyruvate Carboxylase Saccharomyces cerevisiae Sequence Analysis, DNA Suppression, Genetic genetics growth & development genetics |
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| Mesh Heading Relevant : | Genes, Suppressor enzymology genetics metabolism |
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