High level expression of Saccharomyces cerevisiae chitinase (ScCTS1) in Pichia pastoris for degrading chitin

Zhao Youxi, Jiang Huihui, Rao Zhiming, Ji Yizhi, Cheng Yanling, Ma Yanhe

Abstract


Chitin is the second most abundant renewable biopolymer in the world. Chitinases play important roles in the degradation of chitin. Chitinases are produced by different organisms for different purposes, which are widely expressed in the three domains of life, ranging from archaea, bacteria, to fungi, yeasts, plants, insects, and even vertebrates. But there are few reports about Saccharomyces cerevisiae chitinase (ScCTS1). The aim of this study was to realize the high level expression of ScCTS1. The ScCTS1 was cloned into the expression vector pPIC9K. The recombinant plasmid was linearized and transformed into competent Pichia pastoris GS115. After screening by G418 plate, the fermentation conditions were optimized. Ultimately, under the optimal fermentation conditions, ScCTS1 enzymatic activity reached up to 94.6 U/mL. This paper presents the first report on the heterologous expression of a full-length ScCTS1 with considerably high activity. The work will not only make a great stride towards its potential applications in biotechnology, but also facilitate elucidating the precise mechanism of yeast cell division.
Keywords: chitinases, heterologous expression, Saccharomyces cerevisiae, pichia pastoris, biotechnology
DOI: 10.3965/j.ijabe.20150805.2071

Citation: Zhao Y X, Jiang H H, Rao Z M, Ji Y Z, Cheng Y L, Ma Y H. High level expression of Saccharomyces cerevisiae chitinase (ScCTS1) in Pichia pastoris for degrading chitin. Int J Agric & Biol Eng, 2015; 8(5): 142－150.

Keywords


chitinases, heterologous expression, Saccharomyces cerevisiae, pichia pastoris, biotechnology

Full Text:

PDF

References


Yang S Q, Fu X, Yan Q J, Guo Y, Liu Z Q, Jiang Z Q. Cloning, expression, purification and application of a novel chitinase from a thermophilic marine bacterium Paenibacillus barengoltzii. Food Chemistry, 2016; 192: 1041–1048.

Gortari M C, Hours R A. Biotechnological processes for chitin recovery out of crustacean waste. Electronic Journal of Bitechnology, 2013; 16(3): 1–14.

Hartl L, Zach S, Seidl-Seiboth V. Fungal chitinases: diversity, mechanistic properties and biotechnological potential. Appl Microbiol Biotechnol, 2011; 93(2): 533–543.

Bhattacharya D, Nagpure A, Gupta R K. Bacterial chitinases: properties and potential. Crit Rev Biotechnol, 2007; 27 (1): 21–28.

Hou J, Han J, Cai L, Zhou J, Lu Y, Jin C, et al. Characterization of genes for chitin catabolism in Haloferax mediterranei. Appl Microbiol Biotechnol, 2014; 98(3): 1185–1194.

Meng H M, Wang Z X, Meng X Y. Cloning and expression analysis of the chitinase gene Ifu-chit2 from Isaria fumosorosea. Genetics and Molecular Biology, 2015; 38: 381–389.

Jiang C, Huang R F, Song J L, Huang M R, Xu LA. Genomewide analysis of the chitinase gene family in Populus trichocarpa. J Genet, 2013; 92(1): 121–125.

Zhu Q, Arakane Y, Banerjee D, Beeman R W, Kramer K J, Muthukrishnan S. Domain organization and phylogenetic analysis of the chitinase-like family of proteins in three species of insects. Insect Biochem Mol Biol, 2008; 38(4): 452–466.

Adrangi S, Faramarzi M A. From bacteria to human: A journey into the world of chitinases. Biotechnol Adv, 2013; 31(8): 1786–1795.

Li H, Greene L H. Sequence and Structural Analysis of the Chitinase Insertion Domain Reveals Two Conserved Motifs Involved in Chitin-Binding. Plos one, 2010; 5(1): e8654.

Roncero C, Sanchez Y. Cell separation and the maintenance of cell integrity during cytokinesis in yeast: the assembly of a septum. Yeast, 2010; 27(8): 521–30.

Nunezde C G, Francisco F, Davey M R, Cancho S E, Wilson Z A. Conferred resistance to Botrytis cinerea in Lilium by overexpression of the RCH10 chitinase gene. Plant Cell Rep, 2015; 34(7): 1201–1209.

Songsiriritthigul C, Lapboonrueng S, Pechsrichuang P, Pesatcha P, Yamabhai M. Expression and characterization of Bacillus licheniformis chitinase (ChiA), suitable for bioconversion of chitin waste. Bioresource Technol, 2010; 101(11): 4096–4103.

Wang Y J, Yang Q. Cloning and expression of a novel chitinase chi58 from Chaetomium cupreum in Pichia pastoris. Biochem Genet, 2009; 47(7-8): 547–558.

Banani H, Spadaro D, Zhang D P, Matic S, Garibaldi A, Gullino M L. Postharvest application of a novel chitinase cloned from Metschnikowia fructicola and overexpressed in Pichia pastoris to control brown rot of peaches. International Journal of Food Microbiology, 2015; 199(16): 54–61.

Kuranda M J, Robbins P W. Chitinase is required for cell separation during growth of Saccharomyces cerevisiae. J Biol Chem, 1991; 266(29): 19758–19767.

Murphy R A, Power R F G. Expression of Saccharomyces cerevisiae endochitinase in Aspergillus awamori. Biotechnol Lett, 2001; 23(11): 903–906.

Hurtado-Guerrero R, van Aalten D M. Structure of Saccharomyces cerevisiae chitinase 1 and screening-based discovery of potent inhibitors. Chem Biol, 2007; 14(5): 589–599.

Mazan M, Blanco N, Kovacova K, Firakova Z, Rehulka P, Farkaš V, et al. A novel fluorescence assay and catalytic properties of Crh1 and Crh2 yeast cell wall transglycosylases. Biochem J, 2013; 455(3): 307–318.

Hedfalk K. Further advances in the production of membrane proteins in Pichia pastoris. Bioengineered , 2013; 4(6): 363–367.

Ramon A, Marin M. Advances in the production of membrane proteins in Pichia pastoris. Biotechnol J, 2011; 6(6): 700–706.

Fan Y, Zhang Y, Yang X, Pei X, Guo S, Pei Y. Expression of a Beauveria bassiana chitinase (Bbchit1) in Escherichia coli and Pichia pastoris. Protein Expres Purif, 2007: 56(1): 93–99.

Tang W, Li Y, Liu L, Zhang J, Xian H. Characterization and production optimization of a chitinase (Tachil) from Trichoderma asperellum in recombinant Pichia pastoris expression system. Wei Sheng Wu Xue Bao, 2012; 52(3): 345–352. (in Chinese with English abstract)

Goodrick J C, Xu M, Finnegan R, Schilling B M, Schiavi S, Hoppe H, et al. High-level expression and stabilization of recombinant human chitinase produced in a continuous constitutive Pichia pastoris expression system. Biotechnol Bioeng, 2001; 74(6): 492–497.

Xiao X, Zhou Y, Wang F. Isolation and identification of a high-efficient chitin-degrading marine bacterium CBl01 and studies on its chitinases system. Acta Oceanologica Sinica, 2003; 25(1): 138–142. (in Chinese)

Vogl T, Glieder A. Regulation of Pichia pastoris promoters and its consequences for protein production. N Biotechnol, 2013; 30(4): 385–404.

Jahic M, Veide A, Charoenrat T, Teeri T, Enfors S O. Process technology for production and recovery of heterologous proteins with Pichia pastoris. Biotechnol Prog, 2006; 22(6): 1465–1473.




Copyright (c)



2023-2026 Copyright IJABE Editing and Publishing Office