Features & BeneFits Background ligand chemistry covalent Binding single fusion construct more information HaloTag ® Technology Driving mammalian protein expression forward ✓ Covalent binding of interchangeable synthec ligands to fusion tag ✓ Use a single clone for protein purificaon, pull-down, and imaging studies ✓ Higher recovery, purity, funconality and solubility compared to other fusion tags Compared to other tags: • higher capacity than flag® • cleaner than his 6 • no leaching (unlike gst) • more flexible than gfP
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F eat u r es & B e n eF i ts
Background
ligand chemistry
covalent Binding
single fusion construct
more information
HaloTag® TechnologyDriving mammalian protein expression forward
✓ Covalent binding of interchangeable synthetic ligands to fusion tag
✓ Use a single clone for protein purification, pull-down, and imaging studies
✓ Higher recovery, purity, functionality and solubility compared to other fusion tags
Compared to other tags:
• higher capacity than flag®
• cleaner than his6
• no leaching (unlike gst)
• more flexible than gfP
H a l o ta g ® t e c H n o l o g y
features & Benefits
Bac kg ro u n d
ligand chemistry
covalent Binding
single fusion construct
more information
N
T r p 1 0 7
H N A s n 4 1 H
C l
R C A s p 1 0 6
O - O
H N
N H O
C O
G l u 1 3 0
H i s 2 7 2
O H
1 3
2 l
N
T r p 1 0 7
H N A s n 4 1 H
C l
C O O
O H
-O C
O Glu1 3 0
A s p 1 0 6
R
H
P h e 2 7 2
Catalytic process Facilitated bond formation
9755
MA
Origin and optimizationOptimized for mammalian protein expression
dhaaorigin: Rhodococcus rhodochrous Function: forms a covalent intermediate at his272 with chloroalkane substrates by removing cl- upon hydrolysis with water.
Halotag®
optimization: his272Phe mutation allows formation of a stable covalent bond. additional aa changes were made in dhaa to further optimize fusion tag performance.
result: a fusion tag which rapidly forms a covalent attachment to synthetic chloroalkane-based ligands. ideal for protein purification, pull-downs and imaging.
H a l o ta g ® t e c H n o l o g y
features & Benefits
Background
l i g a n d cH e m i st ry
covalent Binding
single fusion construct
more information
HaloTag® ligand chemistry allows interchangeable functionality
• the reactive linker is identical between all halotag® ligands
• move fluidly between protein purification, pull-downs and imaging assays by switching to a new functional group.
Functional group
HaloTag
Proteinof interest
+
Cl
Functional group
HaloTag
Proteinof interest
9756
MA
H a l o ta g ® t e c H n o l o g y
features & Benefits
Background
ligand chemistry
cova l e n t B i n d i n g
single fusion construct
more information
HaloTag® structure and the site of the covalent bond
4787TE
HaloTag® TMRLigand
N-terminus
C-terminus
TMR(functional group)
Chloroalkane(reactive linker)
Phe272
Site of covalent interaction
H a l o ta g ® t e c H n o l o g y
features & Benefits
Background
ligand chemistry
covalent Binding
s i n g l e F u s i o n co n st r u c t
more information
HaloTag® ligandsMany functionalities from a single fusion construct
R
HaloTag®
LigandHaloTag®
Fluorescent ligands
Surfaces
Reactive ligands
9699
MA
oore
Surfac
FFllu
S
solid surfaces• nonmagnetic resin • magnetic beads• customizable arrays
Fluorescent ligands• multiple fluorophores• cell-permeable ligands• cell-impermeable ligands
reactive & capture ligands• introduce novel functionalities
with succinimidyl ester, amine, iodoacetamide and thiol reactive groups
• tag and capture with biotin affinity groups
H a l o ta g ® t e c H n o l o g y
features & Benefits
Background
ligand chemistry
covalent Binding
single fusion construct
m o r e i n Fo r m at i o n
Promega corPoration • 2800 Woods holloW road • madison, Wi 53711-5399 usa • telePhone 608-274-4330 www.promega.com • ©2011 Promega corPoration • all rights reserved • Prices and sPecifications suBject to change Without Prior notice • Printed in usa 2/11 • 19764 • Part #is031
More information
Get started
P r o d u c t s i z e c ata l o g #
Halotag® cloning starter system 1 each g6050
val idated Halotag® human orFs 20µg dna kazusa fhc clones
halotag is a registered trademark of Promega corporation. halolink is a trademark of Promega corporation.
flag is a registered trademark of sigma aldrich.
Protein purification…
Protein interactions…
cellular imaging and labeling…
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