Purification of recombinant α-amylase with immuno-affinity chromatography using monoclonal antibody

Masamichi Kamihira; Masayuki Taniguchi; Shinji Iijima; Takeshi Kobayashi

doi:10.1016/0385-6380(88)90066-0

Purification of recombinant α-amylase with immuno-affinity chromatography using monoclonal antibody

Masamichi Kamihira, Masayuki Taniguchi, Shinji Iijima, Takeshi Kobayashi

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

1 Citation (Scopus)

Abstract

A monoclonal antibody against recombinant thermostable α-amylase produced by Escherichia coli was isolated from serum-free medium and immobilized on Sepharose 4B. The adsorption equilibrium between α-amylase and the immobilized immuno-adsorbent showed a Langmuir type isotherm. The breakthrough curve calculated numerically using the averaged volumetric coefficient coincided well with the experimental data. More than 90% of the activity of bound α-amylase could be recovered by eluting with glycine-HCl buffer (pH 2.5). The elution profile at pH 2.5 became sharper with increasing temperature. By using an immuno-affinity column, the recombinant α-amylase produced by E. coli could be purified homogeneously from crude extract enzyme solution with two-step elution.

Original language	English
Pages (from-to)	625-631
Number of pages	7
Journal	Journal of Fermentation Technology
Volume	66
Issue number	6
DOIs	https://doi.org/10.1016/0385-6380(88)90066-0
Publication status	Published - 1988
Externally published	Yes

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@article{0ee692c2fc944350b73999c6d9431633,

title = "Purification of recombinant α-amylase with immuno-affinity chromatography using monoclonal antibody",

abstract = "A monoclonal antibody against recombinant thermostable α-amylase produced by Escherichia coli was isolated from serum-free medium and immobilized on Sepharose 4B. The adsorption equilibrium between α-amylase and the immobilized immuno-adsorbent showed a Langmuir type isotherm. The breakthrough curve calculated numerically using the averaged volumetric coefficient coincided well with the experimental data. More than 90% of the activity of bound α-amylase could be recovered by eluting with glycine-HCl buffer (pH 2.5). The elution profile at pH 2.5 became sharper with increasing temperature. By using an immuno-affinity column, the recombinant α-amylase produced by E. coli could be purified homogeneously from crude extract enzyme solution with two-step elution.",

author = "Masamichi Kamihira and Masayuki Taniguchi and Shinji Iijima and Takeshi Kobayashi",

note = "Funding Information: This work was supported in part by a Grant-in-Aid for the Encouragement of Young Scientists (No. 63790324) from the Ministry of Education, Science and Culture, Japan.",

year = "1988",

doi = "10.1016/0385-6380(88)90066-0",

language = "English",

volume = "66",

pages = "625--631",

journal = "Journal of Fermentation Technology",

issn = "0385-6380",

publisher = "The Society for Biotechnology, Japan",

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TY - JOUR

T1 - Purification of recombinant α-amylase with immuno-affinity chromatography using monoclonal antibody

AU - Kamihira, Masamichi

AU - Taniguchi, Masayuki

AU - Iijima, Shinji

AU - Kobayashi, Takeshi

N1 - Funding Information: This work was supported in part by a Grant-in-Aid for the Encouragement of Young Scientists (No. 63790324) from the Ministry of Education, Science and Culture, Japan.

PY - 1988

Y1 - 1988

N2 - A monoclonal antibody against recombinant thermostable α-amylase produced by Escherichia coli was isolated from serum-free medium and immobilized on Sepharose 4B. The adsorption equilibrium between α-amylase and the immobilized immuno-adsorbent showed a Langmuir type isotherm. The breakthrough curve calculated numerically using the averaged volumetric coefficient coincided well with the experimental data. More than 90% of the activity of bound α-amylase could be recovered by eluting with glycine-HCl buffer (pH 2.5). The elution profile at pH 2.5 became sharper with increasing temperature. By using an immuno-affinity column, the recombinant α-amylase produced by E. coli could be purified homogeneously from crude extract enzyme solution with two-step elution.

AB - A monoclonal antibody against recombinant thermostable α-amylase produced by Escherichia coli was isolated from serum-free medium and immobilized on Sepharose 4B. The adsorption equilibrium between α-amylase and the immobilized immuno-adsorbent showed a Langmuir type isotherm. The breakthrough curve calculated numerically using the averaged volumetric coefficient coincided well with the experimental data. More than 90% of the activity of bound α-amylase could be recovered by eluting with glycine-HCl buffer (pH 2.5). The elution profile at pH 2.5 became sharper with increasing temperature. By using an immuno-affinity column, the recombinant α-amylase produced by E. coli could be purified homogeneously from crude extract enzyme solution with two-step elution.

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JO - Journal of Fermentation Technology

JF - Journal of Fermentation Technology

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ER -

Purification of recombinant α-amylase with immuno-affinity chromatography using monoclonal antibody

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