Expression and purification of polyhistidine-tagged firefly luciferase in insect cells

A potential alternative for process scale-up

Patrik Michel, Tuula Torkkeli, Matti Karp, Christer Oker-Blom (Corresponding Author)

Research output: Contribution to journalArticleScientificpeer-review

12 Citations (Scopus)

Abstract

The coleopteran firefly, Photinus pyralis, luciferase was produced in lepidopteran Trichoplusia ni insect cells using a baculovirus expression vector. The recombinant protein was equipped with a polyhistidine affinity tag at the carboxyl terminus and purified by immobilized metal-ion affinity chromatography in combination with an expanded bed adsorption system. This approach enabled an efficient, one-step purification protocol of a genetically modified luciferase with properties similar to those of the authentic counterpart. According to light emission measurements, the final yield of highly purified protein was 23 mg l−1 of cell culture. In addition, no specific interaction of interfering substances, such as, ATP, adenylate kinase, nucleoside diphosphokinase, as well as, creatine kinase of the final preparation were identified. Together, the results presented here clearly show that the baculovirus expression system in combination with immobilized metal-ion affinity chromatography is a potential strategy for process scale-up of polyhistidine tagged insect luciferase.
Original languageEnglish
Pages (from-to)49-56
Number of pages8
JournalJournal of Biotechnology
Volume85
Issue number1
DOIs
Publication statusPublished - 2001
MoE publication typeA1 Journal article-refereed

Fingerprint

Firefly Luciferases
Affinity chromatography
Ion chromatography
Baculoviridae
Luciferases
Affinity Chromatography
Purification
Metal ions
Insects
Fireflies
Metals
Nucleoside-Diphosphate Kinase
Ions
Adenylate Kinase
Recombinant proteins
Adenosinetriphosphate
Light emission
Creatine Kinase
Cell culture
Recombinant Proteins

Cite this

Michel, Patrik ; Torkkeli, Tuula ; Karp, Matti ; Oker-Blom, Christer. / Expression and purification of polyhistidine-tagged firefly luciferase in insect cells : A potential alternative for process scale-up. In: Journal of Biotechnology. 2001 ; Vol. 85, No. 1. pp. 49-56.
@article{b81a910231434a7e8c0caff986d3bd52,
title = "Expression and purification of polyhistidine-tagged firefly luciferase in insect cells: A potential alternative for process scale-up",
abstract = "The coleopteran firefly, Photinus pyralis, luciferase was produced in lepidopteran Trichoplusia ni insect cells using a baculovirus expression vector. The recombinant protein was equipped with a polyhistidine affinity tag at the carboxyl terminus and purified by immobilized metal-ion affinity chromatography in combination with an expanded bed adsorption system. This approach enabled an efficient, one-step purification protocol of a genetically modified luciferase with properties similar to those of the authentic counterpart. According to light emission measurements, the final yield of highly purified protein was 23 mg l−1 of cell culture. In addition, no specific interaction of interfering substances, such as, ATP, adenylate kinase, nucleoside diphosphokinase, as well as, creatine kinase of the final preparation were identified. Together, the results presented here clearly show that the baculovirus expression system in combination with immobilized metal-ion affinity chromatography is a potential strategy for process scale-up of polyhistidine tagged insect luciferase.",
author = "Patrik Michel and Tuula Torkkeli and Matti Karp and Christer Oker-Blom",
year = "2001",
doi = "10.1016/S0168-1656(00)00377-1",
language = "English",
volume = "85",
pages = "49--56",
journal = "Journal of Biotechnology",
issn = "0168-1656",
publisher = "Elsevier",
number = "1",

}

Expression and purification of polyhistidine-tagged firefly luciferase in insect cells : A potential alternative for process scale-up. / Michel, Patrik; Torkkeli, Tuula; Karp, Matti; Oker-Blom, Christer (Corresponding Author).

In: Journal of Biotechnology, Vol. 85, No. 1, 2001, p. 49-56.

Research output: Contribution to journalArticleScientificpeer-review

TY - JOUR

T1 - Expression and purification of polyhistidine-tagged firefly luciferase in insect cells

T2 - A potential alternative for process scale-up

AU - Michel, Patrik

AU - Torkkeli, Tuula

AU - Karp, Matti

AU - Oker-Blom, Christer

PY - 2001

Y1 - 2001

N2 - The coleopteran firefly, Photinus pyralis, luciferase was produced in lepidopteran Trichoplusia ni insect cells using a baculovirus expression vector. The recombinant protein was equipped with a polyhistidine affinity tag at the carboxyl terminus and purified by immobilized metal-ion affinity chromatography in combination with an expanded bed adsorption system. This approach enabled an efficient, one-step purification protocol of a genetically modified luciferase with properties similar to those of the authentic counterpart. According to light emission measurements, the final yield of highly purified protein was 23 mg l−1 of cell culture. In addition, no specific interaction of interfering substances, such as, ATP, adenylate kinase, nucleoside diphosphokinase, as well as, creatine kinase of the final preparation were identified. Together, the results presented here clearly show that the baculovirus expression system in combination with immobilized metal-ion affinity chromatography is a potential strategy for process scale-up of polyhistidine tagged insect luciferase.

AB - The coleopteran firefly, Photinus pyralis, luciferase was produced in lepidopteran Trichoplusia ni insect cells using a baculovirus expression vector. The recombinant protein was equipped with a polyhistidine affinity tag at the carboxyl terminus and purified by immobilized metal-ion affinity chromatography in combination with an expanded bed adsorption system. This approach enabled an efficient, one-step purification protocol of a genetically modified luciferase with properties similar to those of the authentic counterpart. According to light emission measurements, the final yield of highly purified protein was 23 mg l−1 of cell culture. In addition, no specific interaction of interfering substances, such as, ATP, adenylate kinase, nucleoside diphosphokinase, as well as, creatine kinase of the final preparation were identified. Together, the results presented here clearly show that the baculovirus expression system in combination with immobilized metal-ion affinity chromatography is a potential strategy for process scale-up of polyhistidine tagged insect luciferase.

U2 - 10.1016/S0168-1656(00)00377-1

DO - 10.1016/S0168-1656(00)00377-1

M3 - Article

VL - 85

SP - 49

EP - 56

JO - Journal of Biotechnology

JF - Journal of Biotechnology

SN - 0168-1656

IS - 1

ER -