EC Number |
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3.1.3.72 | expression in Pichia pastoris |
3.1.3.72 | gene appA or phyS, cloning from library, DNA and amino acid sequence determination and analysis, sequence comparisons and phylogenetic analysis, recombinant expression of extracellular His-tagged enzyme in Escherichia coli strain BL21(DE3) as non-glycosylated protein and of extracellular nontagged enzyme in Pichia pastoris strain GS115(his4) under control of the AOX1 promoter as glycosylated protein |
3.1.3.72 | gene phyAsrl, DNA and amino acid sequence determination and analysis, phyAsrl is constitutively expressed |
3.1.3.72 | recombinant extracellular expression of Lilium longiflorum alkaline phytase in Pichia pastoris strain X33. For analysis of the influence of the native signal sequences on alpha-MF-directed secretion, three expression constructs are generated by inserting three forms of rLlAlp2 downstream of the alpha-MF secretion signal in pPICZaAvectors: 1. full length optimized rLlAlp2 sequence (pPICZaA-op-rLlAlp2), 2. truncated oprLlAlp2 without QKTEL at the C-terminus (pPICZaA-op-LlAlp2DC), and 3. truncated op-rLlAlp2 without the first 19 aa at the N-terminus and without QKTEL at the C-terminus (pPICZaA-oprLlAlp2DNDC). By switching from the strong, tightly regulated promoter PAOX1 to the weaker, constitutive promoter PGAP and optimizing culture conditions, an 8 to 10fold increase in extracellular expression of rLlALP2 is achieved, optimization of culture conditions for enhanced expression, method evaluation, overview |