A constitutive expression system for Pichia pastoris based on the PGK1 promoter

被引：13

作者：

Arruda, Andrelisse ^{[1
]}

Branco Reis, Viviane Castelo ^{[1
]}

Ferreira Batista, Vinicius Daniel ^{[1
]}

Daher, Bruno Sahim ^{[1
]}

Piva, Luiza Cesca ^{[1
]}

De Marco, Janice Lisboa ^{[1
]}

Pepe de Moraes, Lidia Maria ^{[1
]}

Goncalves Torres, Fernando Araripe ^{[1
]}

机构：

[1] Univ Brasilia, Inst Ciencias Biol, Dept Biol Celular, BR-70901900 Brasilia, DF, Brazil

来源：

BIOTECHNOLOGY LETTERS | 2016年 / 38卷 / 03期

关键词：

alpha-Amylase; Constitutive expression; 3-Phosphoglycerate kinase; Pichia pastoris; Recombinant protein production; HETEROLOGOUS PROTEIN-PRODUCTION; FOREIGN GENE-EXPRESSION; YEAST; TRANSFORMANTS; SELECTION; VECTOR; STRAIN;

D O I：

10.1007/s10529-015-2002-2

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

Objectives To develop a new vector for constitutive expression in Pichia pastoris based on the endogenous glycolytic PGK1 promoter. Results P. pastoris plasmids bearing at least 415 bp of PGK1 promoter sequences can be used to drive plasmid integration by addition at this locus without affecting cell growth. Based on this result, a new P. pastoris integrative vector, pPICK2, was constructed bearing some features that facilitate protein production in this yeast: a similar to 620 bp PGK1 promoter fragment with three options of restriction sites for plasmid linearization prior to yeast transformation: a codon-optimized alpha-factor secretion signal, a new polylinker, and the kan marker for vector propagation in bacteria and selection of yeast transformants. Conclusions A new constitutive vector for P. pastoris represents an alternative platform for recombinant protein production and metabolic engineering purposes.

引用

页码：509 / 517

页数：9

共 19 条

[1] A novel kanamycin/G418 resistance marker for direct selection of transformants in Escherichia coli and different yeast species [J].

Agaphonov, Michael ;

Romanova, Nina ;

Choi, Eui-Sung ;

Ter-Avanesyan, Michael .

YEAST, 2010, 27 (04) :189-195

[2] Genetic characterization and construction of an auxotrophic strain of Saccharomyces cerevisiae JP1, a Brazilian industrial yeast strain for bioethanol production [J].

Branco Reis, Viviane Castelo ;

Nicola, Andre Moraes ;

Oliveira Neto, Osmar de Souza ;

Ferreira Batista, Vinicius Daniel ;

Pepe de Moraes, Lidia Maria ;

Goncalves Torres, Fernando Araripe .

JOURNAL OF INDUSTRIAL MICROBIOLOGY & BIOTECHNOLOGY, 2012, 39 (11) :1673-1683

[3] Recombinant protein production in Pichia pastoris under glyceraldehyde-3-phosphate dehydrogenase promoter: From carbon source metabolism to bioreactor operation parameters [J].

Calik, Pinar ;

Ata, Ozge ;

Gunes, Hande ;

Massahi, Aslan ;

Boy, Erdem ;

Keskin, Abdullah ;

Ozturk, Sibel ;

Zerze, Gul H. ;

Ozdamar, Tuncer H. .

BIOCHEMICAL ENGINEERING JOURNAL, 2015, 95 :20-36

[4] New selectable marker/auxotrophic host strain combinations for molecular genetic manipulation of Pichia pastoris [J].

Cereghino, GPL ;

Cereghino, JL ;

Sunga, AJ ;

Johnson, MA ;

Lim, M ;

Gleeson, MAG ;

Cregg, JM .

GENE, 2001, 263 (1-2) :159-169

[5] Expression of heterologous proteins in Pichia pastoris:: a useful experimental tool in protein engineering and production [J].

Daly, R ;

Hearn, MTW .

JOURNAL OF MOLECULAR RECOGNITION, 2005, 18 (02) :119-138

[6] Molecular characterization of the 3-phosphoglycerate kinase gene (PGK1) from the methylotrophic yeast Pichia pastoris [J].

de Almeida, JRM ;

de Moraes, LMP ;

Torres, FAG .

YEAST, 2005, 22 (09) :725-737

[7]

FRAENKEL D.G., 1982, MOL BIOL YEAST SACCH, P1, DOI [10.1016/j.mpsur.2008.12.002, DOI 10.1016/J.MPSUR.2008.12.002]

[8] A NEW METHOD FOR MICRODETERMINATION OF AMYLASE ACTIVITY BY THE USE OF AMYLOSE AS THE SUBSTRATE [J].

FUWA, H .

JOURNAL OF BIOCHEMISTRY, 1954, 41 (05) :583-603

[9] pPCV, a versatile vector for cloning PCR products [J].

Janner, Christiane R. ;

Brito, Ana Livia P. ;

Moraes, Lidia Maria P. ;

Reis, Viviane C. B. ;

Torres, Fernando A. G. .

SPRINGERPLUS, 2013, 2 :1-3

[10] Constitutive expression of barley α-amylase in Pichia pastoris by high-density cell culture [J].

Liu, Z. W. ;

Yin, H. X. ;

Yi, X. P. ;

Zhang, A. L. ;

Luo, J. X. ;

Zhang, T. Y. ;

Fu, C. Y. ;

Zhang, Z. H. ;

Shen, J. C. ;

Chen, L. P. .

MOLECULAR BIOLOGY REPORTS, 2012, 39 (05) :5805-5810

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