Expression and purification of 15 kDa granulysin utilizing an insect cell secretion system

Michael W. Finn; Carol Clayberger; Alan M. Krensky

doi:10.1016/j.pep.2010.07.009

Expression and purification of 15 kDa granulysin utilizing an insect cell secretion system

Michael W. Finn, Carol Clayberger^*, Alan M. Krensky

^*Corresponding author for this work

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

7 Scopus citations

Abstract

Granulysin is an antimicrobial and proinflammatory protein expressed in activated human T cells and natural killer cells. A single mRNA produces the 15 kDa isoform which is then cleaved at the amino and carboxy termini to produce the 9 kDa isoform. Recombinant 9 kDa granulysin has been studied in detail but little is known about the function of the 15 kDa isoform, and no protocol has been published describing expression and purification of this form. Two commercially available preparations of the recombinant 15 kDa granulysin contain tags that may affect function. Here we describe for the first time a method to produce 15 kDa granulysin as a secreted protein from insect cells. The 15 kDa granulysin is purified using a HiTrap Heparin column and a Resource S column. A typical a yield of purified 15 kDa granulysin is 0.6 mg/L of insect cell supernatant.

Original language	English (US)
Pages (from-to)	70-74
Number of pages	5
Journal	Protein Expression and Purification
Volume	75
Issue number	1
DOIs	https://doi.org/10.1016/j.pep.2010.07.009
State	Published - Jan 2011

Keywords

Granulysin
Insect cell expression
Monocytes
NK-cells
Recombinant protein
T cells

ASJC Scopus subject areas

Biotechnology

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10.1016/j.pep.2010.07.009

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title = "Expression and purification of 15 kDa granulysin utilizing an insect cell secretion system",

abstract = "Granulysin is an antimicrobial and proinflammatory protein expressed in activated human T cells and natural killer cells. A single mRNA produces the 15 kDa isoform which is then cleaved at the amino and carboxy termini to produce the 9 kDa isoform. Recombinant 9 kDa granulysin has been studied in detail but little is known about the function of the 15 kDa isoform, and no protocol has been published describing expression and purification of this form. Two commercially available preparations of the recombinant 15 kDa granulysin contain tags that may affect function. Here we describe for the first time a method to produce 15 kDa granulysin as a secreted protein from insect cells. The 15 kDa granulysin is purified using a HiTrap Heparin column and a Resource S column. A typical a yield of purified 15 kDa granulysin is 0.6 mg/L of insect cell supernatant.",

keywords = "Granulysin, Insect cell expression, Monocytes, NK-cells, Recombinant protein, T cells",

author = "Finn, {Michael W.} and Carol Clayberger and Krensky, {Alan M.}",

note = "Funding Information: This work was supported by the Intramural Research Program of the National Institutes of Health , National Cancer Institute , and the Center for Cancer Research . We also thank the members of the Protein Expression Laboratory at NCI Frederick.",

year = "2011",

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doi = "10.1016/j.pep.2010.07.009",

language = "English (US)",

volume = "75",

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journal = "Protein Expression and Purification",

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AU - Finn, Michael W.

AU - Clayberger, Carol

AU - Krensky, Alan M.

N1 - Funding Information: This work was supported by the Intramural Research Program of the National Institutes of Health , National Cancer Institute , and the Center for Cancer Research . We also thank the members of the Protein Expression Laboratory at NCI Frederick.

PY - 2011/1

Y1 - 2011/1

N2 - Granulysin is an antimicrobial and proinflammatory protein expressed in activated human T cells and natural killer cells. A single mRNA produces the 15 kDa isoform which is then cleaved at the amino and carboxy termini to produce the 9 kDa isoform. Recombinant 9 kDa granulysin has been studied in detail but little is known about the function of the 15 kDa isoform, and no protocol has been published describing expression and purification of this form. Two commercially available preparations of the recombinant 15 kDa granulysin contain tags that may affect function. Here we describe for the first time a method to produce 15 kDa granulysin as a secreted protein from insect cells. The 15 kDa granulysin is purified using a HiTrap Heparin column and a Resource S column. A typical a yield of purified 15 kDa granulysin is 0.6 mg/L of insect cell supernatant.

AB - Granulysin is an antimicrobial and proinflammatory protein expressed in activated human T cells and natural killer cells. A single mRNA produces the 15 kDa isoform which is then cleaved at the amino and carboxy termini to produce the 9 kDa isoform. Recombinant 9 kDa granulysin has been studied in detail but little is known about the function of the 15 kDa isoform, and no protocol has been published describing expression and purification of this form. Two commercially available preparations of the recombinant 15 kDa granulysin contain tags that may affect function. Here we describe for the first time a method to produce 15 kDa granulysin as a secreted protein from insect cells. The 15 kDa granulysin is purified using a HiTrap Heparin column and a Resource S column. A typical a yield of purified 15 kDa granulysin is 0.6 mg/L of insect cell supernatant.

KW - Granulysin

KW - Insect cell expression

KW - Monocytes

KW - NK-cells

KW - Recombinant protein

KW - T cells

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JO - Protein Expression and Purification

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Expression and purification of 15 kDa granulysin utilizing an insect cell secretion system

Abstract

Keywords

ASJC Scopus subject areas

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