Comparative Analysis of the Expression Efficiency of the Bacterial Phytase Genes in Pichia pastoris Yeast by the Plate Test

Cyclic AMP dependent protein kinase (PKA) from Pichia pastoris yeast cells was found to be activated by either cAMP or cGMP. Analogs of cAMP such as 8-chloro-cAMP and 8-bromo-cAMP were as potent as cAMP in PKA activation while N6,2'-O-dibutyryl-cAMP did not stimulate the enzyme activity. It was shown that protamine sulfate was almost equally phosphorylated in the presence of 1-2 x 10(-6)M cAMP or cGMP while other substrates such as Kemptide, ribosomal protein S6, were phosphorylated to a lower extent in the presence of cGMP. It was demonstrated that pyruvate kinase is a substrate of PKA which co-purified with the P.pastoris enzyme. Moreover, pyruvate kinase was phosphorylated by PKA in the presence of cAMP and cGMP to comparable levels.

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Increase in the Production of Endo-1,4-β-Xylanase from Paenibacillus brasilensis in Pichia pastoris

Biotekhnologiya ◽

10.21519/0234-2758-2019-35-6-30-38 ◽

2019 ◽

Vol 35 (6) ◽

pp. 30-38 ◽

Cited By ~ 1

Author(s):

L.N. Borshchevskaya ◽

T.L. Gordeeva ◽

S.P. Sineoky

Keyword(s):

Pichia Pastoris ◽

Target Gene ◽

Heterologous Protein ◽

Ministry Of Education ◽

Kurchatov Institute ◽

Xylanase Production ◽

Resource Center ◽

Recombination System ◽

Codon Composition ◽

Pichia Pastoris Yeast

A Pichia pastoris yeast strain producing endo-l,4-β-xylanase from Paenibacillus brasilensis with an activity of 54,400 U/mL after 140 h of fermentation in a laboratory fermenter has been obtained. A number of approaches were used to increase the level of the xylanase production in this strain: optimization of the target gene codon composition, multiple integration of the expression cassette into the recipient strain chromosome using the Cre-lox recombination system, and also improving the heterologous protein folding via the overexpression of the HAC1i gene from Pichia pastoris. xylanase, xylan, Cre-lox system, HAC1p transcriptional activator, multicopy strain, Paenibacillus brasilensis, Pichia pastoris The work was performed with the financial support of the Ministry of Education and Science of Russia (Unique Project Identifier RFMEFI60717X0180) using the Multipurpose Scientific Installation of «All-Russian Collection of Industrial Microorganisms» National Bio-Resource Center, NRC «Kurchatov Institute» - GosNIIgenetika.

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Expression of Aspergillus aculeatus β-mannanase in Pichia pastoris Yeast and Analysis of Industrially Important Properties of Enzyme

Biotekhnologiya ◽

10.21519/0234-2758-2019-35-1-38-44 ◽

2019 ◽

Vol 35 (1) ◽

pp. 38-44

Author(s):

M.N. Lazareva ◽

E.I. Semenko ◽

S.P. Sineoky

Keyword(s):

Pichia Pastoris ◽

Specific Activity ◽

High Specific Activity ◽

Yeast Cells ◽

Aspergillus Aculeatus ◽

Gene Encoding ◽

Pichia Pastoris Yeast ◽

Efficient Expression ◽

Ph Range ◽

First Time

β-Mannanases are enzymes for the industrial application and they can be used, in particular, in the feed industry. The most important requirements for feed enzymes are broad pH range, thermal stability and high specific activity. The efficient expression of the man1 gene encoding Aspergillus aculeatus β-1,4-mannanases in Pichia pastoris yeast cells has been obtained for the first time. The industrially valuable properties of the enzyme were confirmed. The obtained data indicate that the man1 gene from A. aculeatus is potentially useful for the construction of industrial mannanase producers on the basis of the Pichia pastoris yeast. recombinant β-mannanase, Pichia pastoris, Aspergillus aculeatus, overexpression. The work was financially supported by State project №595-00004-18 PR and used the help of the National Bioresource Center - Russian National Collection of Industrial Microorganisms NRC «Kurchatov Institute» - GosNIIgenetika (Moscow, Russia).

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