Improving the purification of NAD+-dependent formate dehydrogenase from Candida methylica

Emel Biçakçi Ordu; Nevin Gül Karagüler

doi:10.1080/10826060701593233

Improving the purification of NAD+-dependent formate dehydrogenase from Candida methylica

Emel Biçakçi Ordu, Nevin Gül Karagüler^*

^*Corresponding author for this work

Department of Molecular Biology and Genetics

Research output: Contribution to journal › Article › peer-review

14 Citations (Scopus)

Abstract

The Candida methylica (cm) recombinant wild type formate dehydrogenase (FDH) gene has been cloned into the pQE-2 TAGZyme expression vector and the 6xHis-tagged FDH gene has been overexpressed in JM105 cells to purify the FDH protein more efficiently, by the use of exopeptidases, TAGZyme Purification System, which has allowed the complete removal of the small N-terminal His-tag. After the purification procedure, 1.2 mg/mL cmFDH protein of >95% purity was obtained. The kinetic parameters of cmFDH have been determined by observing the oxidation of the nicotinamide coenzyme at 340 nm. The results have also been compared to the yield of standard vs. affinity purification of FDH.

Original language	English
Pages (from-to)	333-341
Number of pages	9
Journal	Preparative Biochemistry and Biotechnology
Volume	37
Issue number	4
DOIs	https://doi.org/10.1080/10826060701593233
Publication status	Published - Oct 2007

Keywords

Candida methylica
Formate dehydrogenase
NAD
Purification

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10.1080/10826060701593233

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title = "Improving the purification of NAD+-dependent formate dehydrogenase from Candida methylica",

abstract = "The Candida methylica (cm) recombinant wild type formate dehydrogenase (FDH) gene has been cloned into the pQE-2 TAGZyme expression vector and the 6xHis-tagged FDH gene has been overexpressed in JM105 cells to purify the FDH protein more efficiently, by the use of exopeptidases, TAGZyme Purification System, which has allowed the complete removal of the small N-terminal His-tag. After the purification procedure, 1.2 mg/mL cmFDH protein of >95% purity was obtained. The kinetic parameters of cmFDH have been determined by observing the oxidation of the nicotinamide coenzyme at 340 nm. The results have also been compared to the yield of standard vs. affinity purification of FDH.",

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AU - Karagüler, Nevin Gül

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AB - The Candida methylica (cm) recombinant wild type formate dehydrogenase (FDH) gene has been cloned into the pQE-2 TAGZyme expression vector and the 6xHis-tagged FDH gene has been overexpressed in JM105 cells to purify the FDH protein more efficiently, by the use of exopeptidases, TAGZyme Purification System, which has allowed the complete removal of the small N-terminal His-tag. After the purification procedure, 1.2 mg/mL cmFDH protein of >95% purity was obtained. The kinetic parameters of cmFDH have been determined by observing the oxidation of the nicotinamide coenzyme at 340 nm. The results have also been compared to the yield of standard vs. affinity purification of FDH.

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Improving the purification of NAD+-dependent formate dehydrogenase from Candida methylica

Abstract

Keywords

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