Difference between revisions of "Ligand Binding"

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[[Catalog|< Back to Catalog]]
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=Overview=
 
=Overview=
  
 
=Protein binding=
 
=Protein binding=
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==Antikörper==
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==Antibodies==
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==Affibody==
 
==Affibody==
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Affibody Z-EGFR-1907
 
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()
 
  
 
Affibodies are small (6 kDa), soluble high-affinity proteins. They are derived from the IgG-binding B domain of the Staphylococcal protein A, which was engineered to specifically bind to certain peptides or proteins. This so-called Z domain consists of an antiparallel three-helix bundle and is advantageous due to its proteolytic and thermodynamic stability, its good folding properties and the ease of production via recombinant bacteria (Nord et al., 1997). Affibodies can be used for example for tumor targeting (Wikman et al., 2004) and diagnostic imaging applications (Orlova et al., 2006; Orlova et al., 2007). The ZEGFR:1907 Affibody was engineered to specifically bind the EGF receptor with an affinity determined to be KD = 2.8 nM (Friedman et al., 2008). The EGF receptor is overexpressed in certain types of tumors, e.g. in breast (Walker & Dearing, 1999), lung (Hirsch et al., 2003) and bladder (Colquhoun & Mellon, 2002) carcinomas, and is therefore a suitable target for cancer imaging or therapeutic applications. Because of their good tumor uptake, and their property to become internalized into the target cells with an efficiency of 19 – 24% within one hour – compared to 45% of the natural ligand EGF - the ZEGFR:1907 Affibody was chosen for therapeutic applications by the Freiburg iGEM Team 2010 (Friedman et al., 2008; Göstring et al., 2010).
 
Affibodies are small (6 kDa), soluble high-affinity proteins. They are derived from the IgG-binding B domain of the Staphylococcal protein A, which was engineered to specifically bind to certain peptides or proteins. This so-called Z domain consists of an antiparallel three-helix bundle and is advantageous due to its proteolytic and thermodynamic stability, its good folding properties and the ease of production via recombinant bacteria (Nord et al., 1997). Affibodies can be used for example for tumor targeting (Wikman et al., 2004) and diagnostic imaging applications (Orlova et al., 2006; Orlova et al., 2007). The ZEGFR:1907 Affibody was engineered to specifically bind the EGF receptor with an affinity determined to be KD = 2.8 nM (Friedman et al., 2008). The EGF receptor is overexpressed in certain types of tumors, e.g. in breast (Walker & Dearing, 1999), lung (Hirsch et al., 2003) and bladder (Colquhoun & Mellon, 2002) carcinomas, and is therefore a suitable target for cancer imaging or therapeutic applications. Because of their good tumor uptake, and their property to become internalized into the target cells with an efficiency of 19 – 24% within one hour – compared to 45% of the natural ligand EGF - the ZEGFR:1907 Affibody was chosen for therapeutic applications by the Freiburg iGEM Team 2010 (Friedman et al., 2008; Göstring et al., 2010).
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{|cellspacing="0" border="1"
 
{|cellspacing="0" border="1"
 
|Name
 
|Name
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|BioBrick RFC10
 
|BioBrick RFC10
 
|BioBrick RFC25
 
|BioBrick RFC25
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|Team
 
|Reference
 
|Reference
 
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|<partinfo>BBa_K404302</partinfo>
 
|<partinfo>BBa_K404302</partinfo>
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|[http://2010.igem.org/Team:Freiburg_Bioware Freiburg 2010]
 
|
 
|
 
|}
 
|}
  
 
==Darpin==
 
==Darpin==
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DARPin-E01
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Natural protein ankyrin repeat (AR) molecules are motifs that can be found commonly in proteins (Bork 1993). These motifs mediate protein-protein interactions suggesting that AR proteins can be used for designing new binding molecules. Design of structural scaffolds with consensus regions and randomized positions of interacting residues leads to improved biophysical characteristics of targeting molecules (Binz et al. 2003) (Kohl et al. 2003).
 
Natural protein ankyrin repeat (AR) molecules are motifs that can be found commonly in proteins (Bork 1993). These motifs mediate protein-protein interactions suggesting that AR proteins can be used for designing new binding molecules. Design of structural scaffolds with consensus regions and randomized positions of interacting residues leads to improved biophysical characteristics of targeting molecules (Binz et al. 2003) (Kohl et al. 2003).
  
 
The repetitive nature of the ankyrin proteins allows modifications in their variable and modular binding surface. Therefore, consensus sequences of natural ankyrin proteins have been used to design novel and stable scaffolds for binding proteins.
 
The repetitive nature of the ankyrin proteins allows modifications in their variable and modular binding surface. Therefore, consensus sequences of natural ankyrin proteins have been used to design novel and stable scaffolds for binding proteins.
 
Designed Ankyrin Repeat Proteins (DARPins) are well expressed, monomeric in solution, thermodynamically stable and have the ability to fold fast. In the publication of (Steiner et al. 2008) screening libraries were created by using the signal recognition particle (SRP) translocation pathway for phage display. The selected DARPin E_01 has very high affinities to the target protein ErbB1 and can be used as a potential targeting molecule for our approach by fusing the DARPin to N-terminal VP proteins. Our designed ankyrin repeat protein consists of three internal binding repeats and the C-and N-terminal capping repeats. Each internal repeat module comprises one beta-turn and two hydrophobic alpha helices. The potential interaction residues are located in the beta-turn and the first alpha helix of the AR-proteins.  
 
Designed Ankyrin Repeat Proteins (DARPins) are well expressed, monomeric in solution, thermodynamically stable and have the ability to fold fast. In the publication of (Steiner et al. 2008) screening libraries were created by using the signal recognition particle (SRP) translocation pathway for phage display. The selected DARPin E_01 has very high affinities to the target protein ErbB1 and can be used as a potential targeting molecule for our approach by fusing the DARPin to N-terminal VP proteins. Our designed ankyrin repeat protein consists of three internal binding repeats and the C-and N-terminal capping repeats. Each internal repeat module comprises one beta-turn and two hydrophobic alpha helices. The potential interaction residues are located in the beta-turn and the first alpha helix of the AR-proteins.  
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<Partinfo>BBa_K404314</partinfo>
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{|cellspacing="0" border="1"
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|Name
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|Target
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|Amino acids
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|Mol. mass [kDa]
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|BioBrick RFC10
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|BioBrick RFC25
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|Team
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|Reference
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|-
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|DARPin-E01
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|HER2
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|
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|
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|
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|<partinfo>BBa_K404302</partinfo>
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|[http://2010.igem.org/Team:Freiburg_Bioware Freiburg 2010]
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|
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|}
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==Anticalin==
 
==Anticalin==
 
The Anticalin technology was developed by Prof. Dr. A. Skerra (TU Munich, Germany) that are based on lipocalins
 
The Anticalin technology was developed by Prof. Dr. A. Skerra (TU Munich, Germany) that are based on lipocalins
 
The anticalins availible as BioBricks are the digoxigenin binding anticalin DigA (<partinfo>BBa_K243003</partinfo>) and the fluorescein binding anticalin FluA (<partinfo>BBa_K157004</partinfo>).
 
The anticalins availible as BioBricks are the digoxigenin binding anticalin DigA (<partinfo>BBa_K243003</partinfo>) and the fluorescein binding anticalin FluA (<partinfo>BBa_K157004</partinfo>).
  
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=Small molecule binding=
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Fluoreszein
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{|cellspacing="0" border="1"
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Digitonin
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|Name
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|Target
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|Amino acids
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|Mol. mass [kDa]
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|BioBrick RFC10
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|BioBrick RFC25
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|Team
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|Reference
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|-
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|DigA
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|Digoxigenin
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|
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|
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|
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|<partinfo>BBa_K243003</partinfo>
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|[http://2009.igem.org/Team:Freiburg_bioware Freiburg 2009]
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|
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|-
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|FluA
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|Fluorescein
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|
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|
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|
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|<partinfo>BBa_K157004</partinfo>
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|[http://2008.igem.org/Team:Freiburg Freiburg 2008]
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|
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|}
  
 
=Sugar binding=
 
=Sugar binding=
 
*Cellulose Binding
 
*Cellulose Binding
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=Biotin binding=

Latest revision as of 14:55, 16 March 2013

< Back to Catalog


Overview

Protein binding

Antibodies

Affibody

Affibodies are small (6 kDa), soluble high-affinity proteins. They are derived from the IgG-binding B domain of the Staphylococcal protein A, which was engineered to specifically bind to certain peptides or proteins. This so-called Z domain consists of an antiparallel three-helix bundle and is advantageous due to its proteolytic and thermodynamic stability, its good folding properties and the ease of production via recombinant bacteria (Nord et al., 1997). Affibodies can be used for example for tumor targeting (Wikman et al., 2004) and diagnostic imaging applications (Orlova et al., 2006; Orlova et al., 2007). The ZEGFR:1907 Affibody was engineered to specifically bind the EGF receptor with an affinity determined to be KD = 2.8 nM (Friedman et al., 2008). The EGF receptor is overexpressed in certain types of tumors, e.g. in breast (Walker & Dearing, 1999), lung (Hirsch et al., 2003) and bladder (Colquhoun & Mellon, 2002) carcinomas, and is therefore a suitable target for cancer imaging or therapeutic applications. Because of their good tumor uptake, and their property to become internalized into the target cells with an efficiency of 19 – 24% within one hour – compared to 45% of the natural ligand EGF - the ZEGFR:1907 Affibody was chosen for therapeutic applications by the Freiburg iGEM Team 2010 (Friedman et al., 2008; Göstring et al., 2010).


Name Target Amino acids Mol. mass [kDa] BioBrick RFC10 BioBrick RFC25 Team Reference
Z-EGFR-1907 HER2 BBa_K404302 [http://2010.igem.org/Team:Freiburg_Bioware Freiburg 2010]

Darpin

Natural protein ankyrin repeat (AR) molecules are motifs that can be found commonly in proteins (Bork 1993). These motifs mediate protein-protein interactions suggesting that AR proteins can be used for designing new binding molecules. Design of structural scaffolds with consensus regions and randomized positions of interacting residues leads to improved biophysical characteristics of targeting molecules (Binz et al. 2003) (Kohl et al. 2003).

The repetitive nature of the ankyrin proteins allows modifications in their variable and modular binding surface. Therefore, consensus sequences of natural ankyrin proteins have been used to design novel and stable scaffolds for binding proteins. Designed Ankyrin Repeat Proteins (DARPins) are well expressed, monomeric in solution, thermodynamically stable and have the ability to fold fast. In the publication of (Steiner et al. 2008) screening libraries were created by using the signal recognition particle (SRP) translocation pathway for phage display. The selected DARPin E_01 has very high affinities to the target protein ErbB1 and can be used as a potential targeting molecule for our approach by fusing the DARPin to N-terminal VP proteins. Our designed ankyrin repeat protein consists of three internal binding repeats and the C-and N-terminal capping repeats. Each internal repeat module comprises one beta-turn and two hydrophobic alpha helices. The potential interaction residues are located in the beta-turn and the first alpha helix of the AR-proteins.

Name Target Amino acids Mol. mass [kDa] BioBrick RFC10 BioBrick RFC25 Team Reference
DARPin-E01 HER2 BBa_K404302 [http://2010.igem.org/Team:Freiburg_Bioware Freiburg 2010]

Anticalin

The Anticalin technology was developed by Prof. Dr. A. Skerra (TU Munich, Germany) that are based on lipocalins The anticalins availible as BioBricks are the digoxigenin binding anticalin DigA (BBa_K243003) and the fluorescein binding anticalin FluA (BBa_K157004).


Name Target Amino acids Mol. mass [kDa] BioBrick RFC10 BioBrick RFC25 Team Reference
DigA Digoxigenin BBa_K243003 [http://2009.igem.org/Team:Freiburg_bioware Freiburg 2009]
FluA Fluorescein BBa_K157004 [http://2008.igem.org/Team:Freiburg Freiburg 2008]

Sugar binding

  • Cellulose Binding

Biotin binding