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Updated: Jun 10, 2026
A High Yield and Cost-efficient Expression System of Human Granzymes in Mammalian Cells
Published on: June 10, 2015
Expression and purification of 15 kDa granulysin utilizing an insect cell secretion system
Michael W Finn1, Carol Clayberger, Alan M Krensky
1Laboratory of Cellular and Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, 37 Convent Drive, Room 2016, Bethesda, MD 20892-4256, USA.
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.
Insights
This study presents a novel method for producing and purifying the 15 kDa granulysin protein from insect cells. This advancement facilitates further research into granulysin
Area of Science:
- Immunology
- Protein Biochemistry
Background:
- Granulysin is an antimicrobial and proinflammatory protein found in T cells and natural killer cells.
- The 15 kDa granulysin isoform's function is largely unknown, unlike the well-studied 9 kDa isoform.
- Existing recombinant 15 kDa granulysin preparations may have function-altering tags.
Purpose of the Study:
- To establish a protocol for expressing and purifying the 15 kDa granulysin isoform.
- To produce 15 kDa granulysin as a secreted protein in insect cells.
- To provide a tag-free recombinant 15 kDa granulysin for functional studies.
Main Methods:
- Expression of 15 kDa granulysin in insect cells.
- Purification using HiTrap Heparin and Resource S chromatography.
- Quantification of protein yield.
Main Results:
- Successful production of secreted 15 kDa granulysin from insect cells.
- Purification yielded a tag-free 15 kDa granulysin.
- Typical yield of 0.6 mg/L of insect cell supernatant.
Conclusions:
- A novel and effective method for producing and purifying 15 kDa granulysin has been developed.
- This protocol provides a valuable tool for studying the function of the 15 kDa granulysin isoform.
- The tag-free recombinant protein is suitable for detailed functional analysis.
