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CHOOSE THE CONVENIENT WAY TO MOVE
FROM GENES TO FUNCTIONAL PROTEINS –
THE GST GENE FUSION SYSTEM

18-1159-30
Edition AA
 In Proteomics, speed and
“Spend some time to save time

GST GENE FUSION SYSTEM

KEEPS YOU ON THE FAST TRACK
When working with fusion proteins, essential factors include purity, speed,
yield, preservation of protein structure, and experimental reproducibility.
Glutathione S-Transferase (GST) Gene Fusion System provides solutions
to these factors through an integrated system for the expression, purifica-
tion and detection of fusion proteins produced in E. coli.
GST Gene Fusion System consists of three key components:
pGEX plasmid vectors, GST Purification Modules, and
GST Detection Modules.
Companion products such as site-specific proteases, purification
products and instruments complement the range.There are many
reasons to choose the GST Gene Fusion System, from faster
methodology to higher yields and purity of functional proteins.

• Speed counts
Research today is competitive and in the race for results in
proteomics, a fast methodology that delivers reliable results is
always advantageous. One-step GST tag removal and efficient
protein purification will save you both time and effort.

• “Protein friendly”
What is the point of spending weeks producing fusion proteins if
the purification process damages them? pGEX-6P PreScissionTM
Protease vectors offer GST tag removal and protein purification at
2 4 °C for improved protein stability.
accuracy matter
– can you afford not to read on?”

• Increased yield and purity

A minimal number of steps and highly specific affinity purification
mean higher yields and purity.

• Single source for GST expertise

Amersham Biosciences offers a complete solution for working with
GST fusion proteins. From everything you need for success on a
lab scale to large-scale purification, Amersham Biosciences offers
the reagents, equipment, and expertise designed to meet your
needs – see www.amershambiosciences.com for details.

Amersham Biosciences GST products provide scientists

with the freedom to do experiments from gene to
function to structure in one complementary system.
Dr. Darcy Birse, Department of Biochemistry and Biophysics, Stockholm University
 Good research starts at
pGEX VECTORS ENSURE HIGH YIELDS
OF FUSION PROTEINS
GST Gene Fusion System incorporates pGEX plasmids for inducible,
high-level intracellular expression of genes or gene-fragments as
fusions with Schistosoma japonicum GST. Expression in E. coli yields
fusion proteins in the cytoplasm with the GST moiety at the amino
terminus and the protein of interest at the carboxyl terminus.
Ten pGEX vectors are available in three series – make your choice
based on reading frame and the protease that you want to use
downstream.

4
the beginning

pGEX fusion vectors offer:

• A tac promoter for chemically-inducible, high-level expression.
• An internal lacIq gene so you can use any E. coli host. A
lyophilized culture of E. coli BL21 is supplied with each vector.
This strain is deficient in OmpT and Lon proteases, which
minimizes proteolytic degradation of fusion proteins.
• Choice of protease recognition sites for cleaving the target
protein from the fusion product.
pGEX-6P PreScission Protease vectors offer the most efficient
method for cleaving and purifying GST fusion proteins. Site-specific
cleavage is performed with simultaneous immobilization of the
protease on the column.The protease and GST tag are removed
in a single step.
Other vectors available are pGEX-5X Factor Xa vectors and
pGEX-4T Thrombin vectors. Also available is pGEX-2TK, which
allows the detection of expressed proteins by direct labelling of
fusion products in vitro.
For detailed methodologies, see our technical handbooks listed on
page 10.
5
 Why settle for less
CHOOSE THE PURIFICATION FORMAT
THAT FITS YOUR SCALE
GST purification features Glutathione SepharoseTM affinity media for rapid,
mild affinity purification of GST fusion proteins.
Fusion proteins can be purified to greater than 90% in a single step using mild,
non-denaturing elution conditions with 10 mM reduced glutathione at pH 8.0
that preserves protein antigenicity and function.
You can easily scale up from microgram to hundreds of milligram yields.

MicroSpin GST Purification Module

MicroSpinTM GST Purification Module comprises 50 prepacked Glutathione
Sepharose 4B columns in a convenient MicroSpin format. Each column, with
its 50 µl bed volume, is sufficient to purify up to 400 µg of recombinant GST.
By using MicroPlexTM 24 Vacuum, you can increase the throughput of your
purification process. Up to 48 samples can be handled simultaneously.

GSTrap FF
GSTrapTM FF columns are prepacked 1 ml and 5 ml HiTrapTM columns
containing Glutathione Sepharose 4 Fast Flow with a binding capacity of
10–12 mg GST/ml gel.

GSTPrep FF 16/10
GSTPrepTM FF 16/10 columns are prepacked 20 ml HiPrepTM columns
containing Glutathione Sepharose 4 Fast Flow for preparative purifications of
GST fusion proteins. Each column binds more than 200 mg of fusion protein.

Glutathione Sepharose 4 Fast Flow

Glutathione Sepharose 4 Fast Flow is supplied as pre-swollen bulk medium,
and is an excellent choice for scaling up purifications.

6
than 90% purity?
ON-COLUMN CLEAVAGE WITH
PRESCISSION PROTEASE
Why not purify the target protein at the same time as
removing the GST tag? On-column cleavage is generally
recommended as the method of choice since many
contaminants can be washed out and the cleaved
target protein eluted with a higher level of purity.
With the pGEX-6P vectors, you can use PreScission
Protease to cleave the GST tag from the fusion
protein. PreScission Protease is a fusion protein of
GST and human rhinovirus 3C protease.
• PreScission Protease works best at 4 °C, unlike
other proteases that require temperatures closer
to 20 °C. A lower temperature means increased
protein stability.
• The amino acid sequence recognized by PreScission
Protease is synthetic.This means that the enzyme
will not cleave your target protein.
• Unlike other proteases, once the GST tag is cleaved
off, only a glycine-proline group remains at the
amino-terminal end.This group has virtually no
functional or conformational effect on your protein
(the actual number of non-native amino acids at the
amino terminus will depend on which restriction
site was used for cloning).
• Only the target protein is eluted from the column
while bound to Glutathione Sepharose. GST and
PreScission Protease remain bound to the media.
You can expect purity levels in excess of 90%.

For detailed methodologies, see our technical hand-

books listed on page 10.
 Spend less
time looking
GST 96-WELL DETECTION
MODULE FOR
HIGH-THROUGHPUT SCREENING
GST 96-Well Detection Module permits rapid, sensitive determination
of GST fusion proteins in a variety of samples.
Clarified lysates or intermediate purification fractions can be applied
directly into the wells of GST 96-Well Detection Plates. GST fusion
proteins are captured by specific binding to anti-GST antibody that is
immobilized on the walls of each well.
• Detect captured fusion proteins with the provided horseradish
peroxidase conjugated anti-GST antibody or with an antibody
specific for the target protein.
• Preblocked plates prevent non-specific binding for highly sensitive
8 results with low background.
• Versatile format provides 12 strips of eight wells for processing
8–96 samples at one time. Unused strips can be safely stored until
needed.
Reagents are also available for performing Western blot analysis using
anti-GST antibody.This technique uses only 5–10 µl of sample and is
highly specific.The method also provides information on the size of
the protein.

For detailed methodologies, see our technical handbooks listed on

page 10.
 Fusions without the
confusion
CHOOSE THE GST GENE FUSION SYSTEM
TO MAKE YOUR RESEARCH MORE EFFECTIVE
Take away the worry and effort from the production of purifying GST fusion proteins. When you want to
fusion proteins and focus on the important parts of scale up your production, we have media and
your research. columns to suit a wide range of vectors, purification
media and equipment together with at sensitive
• Optimized and standardized, around the world
applications. We also offer handbooks to help you in
The components of GST Gene Fusion System work
your research.
together, using a standardized methodology.This
means less work in devising experimental protocols. • The right choice
And if you are part of a consortium, you can obtain GST Gene Fusion System is the right choice for
and use the same vector constructs methodology producing and purifying fusion proteins. A wide
and equipment, and rely on getting comparable range of vectors, various purification systems and a
results, anywhere around the world. sensitive detection system all mean that you can
obtain large yields of high-quality target proteins
• All from one
with minimum difficulty.
Amersham Biosciences supplies all the reagents and
instruments you need for producing, detecting and

Find out more about the methodology

Recombinant Protein Handbook (Code No. 18-1142-75)
Covers topics including GST and His fusion proteins, protein amplification, extraction, purifi-
cation, handling of inclusion bodies, methodology, applications, and technical information.

GST Gene Fusion Handbook (Code No. 18-1157-58)

Covers topics including cloning, monitoring, purification, detection, and applications of GST
10 fusion proteins.
Ordering information
Product Quantity Code No.

Cloning and expression

pGEX-6P-1 25 µg 27-4597-01
pGEX-6P-2 25 µg 27-4598-01
pGEX-6P-3 25 µg 27-4599-01
pGEX-5X-1 25 µg 27-4584-01
pGEX-5X-2 25 µg 27-4585-01
pGEX-5X-3 25 µg 27-4586-01
pGEX-4T-1 25 µg 27-4580-01
pGEX-4T-2 25 µg 27-4581-01
pGEX-4T-3 25 µg 27-4583-01
pGEX-2TK 25 µg 27-4587-01
E. coli BL21
(All pGEX vectors include E. coli B21) 1 vial 27-1542-01

Purification
Prepacked columns and media
GST MicroSpin Purification Module 50 purifications 27-4570-03
GSTrap FF 2x1 ml 17-5130-02
5x1 ml 17-5130-01
1x5 ml 17-5131-01
GSTPrep FF 16/10 1x20 ml 17-5234-01
Glutathione Sepharose 4 Fast Flow 25 ml 17-5132-01
100 ml 17-5132-02
500 ml 17-5132-03
Glutathione Sepharose 4B 10 ml 17-0756-01
100 ml 27-4574-01
300 ml 17-0756-04

Removal of trypsin-like serine proteases

HiTrap Benzamidine FF (high sub) 2x1 ml 17-5143-02
5x1 ml 17-5143-01
1x5 ml 17-5144-01
Benzamidine Sepharose 4
Fast Flow (high sub) 25 ml 17-5123-10

Detection
GST 96-Well Detection Module 96 reactions 27-4592-01
GST Detection Module 50 reactions 27-4590-01
Anti-GST antibody 0.5 ml 27-4577-01

Cleavage enzymes
PreScission Protease 500 units 27-0843-01
Factor Xa 400 units 27-0849-01
Thrombin 500 units 27-0846-01

Equipment
MicroPlex 24 Vacuum 1 system 27-3567-01

11
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HiTrap, GSTPrep, GSTrap, HiPrep, MicroPlex, MicroSpin, PreScission,

and Sepharose are trademarks of the Amersham Biosciences Limited.

Amersham and Amersham Biosciences are trademarks of Amersham plc.

Licensing Information
All materials for research use only. Not for diagnostic or therapeutic purposes.
A license for commercial use of pGEX vectors must be obtained from AMRAD Corporation Ltd., 17-27 Cotham Road, Kew,Victoria 3101, Australia.
PreScission Protease is licensed from the University of Singapore, 10 Kent Ridge Crescent, Singapore 0511.
PreScission Protease is produced under commercial license from AMRAD Corporation Ltd., 17-27 Cotham Road, Kew,Victoria 3101, Australia.

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