Cloning and heterologous expression of Laccase in pichia pastoris and determination some of biochemical properties

Authors

  • Nematzadeh, GhorbanAli Genetic & Agricultural Biotechnology Institute of Tabarestan(GABIT). Sari Agricultural Sciences & Natural Resources University (SANRU)
  • Soleimani, Elham Genetic & Agricultural Biotechnology Institute of Tabarestan(GABIT). Sari Agricultural Sciences & Natural Resources University (SANRU)
Abstract:

Laccase (EC 1.10.3.2) are multi-copper oxidase which catalyze the oxidation aromatic and non- aromatic compounds with electron reduction of molecular oxygen to water. Nucleotide sequence of laccase (accession number : ) was optimized according codon preference of Pichia pastoris. Gene was synthesized and cloned into pPICZalpha A. laccase under control of AOX1 promoter was transformed to P.pastoris by electroporation and integrated to P.pastoris genome. Methanol concentration, copper salts concentration and temperature were varied in order to enhance laccase expression in the heterologous system. Optimal fermentation condition for laccase production in shake flask cultivation using BMGY medium were obtained: presence of 0.4 mM Cu+2, culture temperature 25°C, 0.8% methanol added into culture every 24 h. the volumetric activity was achieved 9600 U/L after 12 day culture in fernbach flask. The biochemical properties of recombinant laccase and kinetic parameters were studied. The laccase activity was optimal at pH 4 and 40°C. Its Km value was 0.089 mM For ABTS [2, 2´azino-bis (3-ethylbenzthiazoline-6-sulphoric acid)]. The result of this study was showed Pichia pastoris can be useful tool for heterologous laccase expression in order to use in industrial application such as: waste water treatment, biosensor & etc.

Upgrade to premium to download articles

Sign up to access the full text

Already have an account?login

similar resources

determination of some physical and mechanical properties red bean

چکیده: در این تحقیق، برخی خواص فیزیکی و مکانیکی لوبیا قرمز به-صورت تابعی از محتوی رطوبت بررسی شد. نتایج نشان داد که رطوبت بر خواص فیزیکی لوبیا قرمز شامل طول، عرض، ضخامت، قطر متوسط هندسی، قطر متوسط حسابی، سطح تصویر شده، حجم، چگالی توده، تخلخل، وزن هزار دانه و زاویه ی استقرار استاتیکی در سطح احتمال 1 درصد اثر معنی داری دارد. به طوری که با افزایش رطوبت از 54/7 به 12 درصد بر پایه خشک طول، عرض، ضخام...

15 صفحه اول

some properties of fuzzy hilbert spaces and norm of operators

in this thesis, at first we investigate the bounded inverse theorem on fuzzy normed linear spaces and study the set of all compact operators on these spaces. then we introduce the notions of fuzzy boundedness and investigate a new norm operators and the relationship between continuity and boundedness. and, we show that the space of all fuzzy bounded operators is complete. finally, we define...

15 صفحه اول

Heterologous Expression of Bovine Prochymosin in Pichia pastoris GS115

Objectives: In present research we evaluate the expression of this critical enzyme in a eukaryotic system for future use in cheese industry. Materials and Methods: We have cloned bovine prochymosin gene in methylotrophic yeast, P. pastoris, using pPIC9K as an expression vector. The recombinant plasmid was transformed into the host by electroporation, and it was expre...

full text

Cloning, sequence analysis, expression of Cyathus bulleri laccase in Pichia pastoris and characterization of recombinant laccase

BACKGROUND Laccases are blue multi-copper oxidases and catalyze the oxidation of phenolic and non-phenolic compounds. There is considerable interest in using these enzymes for dye degradation as well as for synthesis of aromatic compounds. Laccases are produced at relatively low levels and, sometimes, as isozymes in the native fungi. The investigation of properties of individual enzymes therefo...

full text

Molecular cloning of a laccase isozyme gene from Pleurotus sajor-caju and expression in the heterologous Pichia pastoris host.

The Psc lac4 gene from Pleurotus sajor-caju has been cloned and expressed in the heterologous host Pichia pastoris, under the control of the AOX1 methanol inducible promoter. The native Ple. sajor-caju laccase signal sequence was effective in directing the secretion of lac4 expressed in Pic. pastoris. The control of media pH and temperature was found to be important in obtaining sufficient quan...

full text

My Resources

Save resource for easier access later

Save to my library Already added to my library

{@ msg_add @}


Journal title

volume 8  issue 2

pages  121- 131

publication date 2019-11

By following a journal you will be notified via email when a new issue of this journal is published.

Keywords

No Keywords

Hosted on Doprax cloud platform doprax.com

copyright © 2015-2023