Difference between revisions of "Part:BBa K4380000:Design"

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===Design===
 
===Design===
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Design
 
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Vilnius-Lithuania Igem 2022 team used this part as a novel way for peptide immobilization. The team was working to create an easily accessible nanoplastic detection tool, using peptides, whose interaction with nanoplastic particles would lead to an easily interpretable response. The system itself focused on smaller protein molecules, peptides, which are modified to acquire the ability to connect to the surface of synthetic polymers – plastics. The detection system works when peptides and nanoplastic particles combine and form a "sandwich" complex - one nanoplastic particle is surrounded by two peptides, attached to their respective protein. The sandwich complex consisted of two main parts – one is peptide bound to a fluorescent protein, other peptide immobilized on cellulose membrane by cellulose binding domain.
 
  
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Vilnius-Lithuania Igem's 2022 team used this part as a novel way for peptide immobilization. The team was working to create an easily accessible nanoplastic detection tool, using peptides, whose interaction with nanoplastic particles would lead to an easily interpretable response. The system itself focused on smaller protein molecules, peptides, which are modified to acquire the ability to connect to the surface of synthetic polymers – plastics. The detection system works when peptides and nanoplastic particles combine and form a "sandwich" complex - one nanoplastic particle is surrounded by two peptides, attached to their respective protein. The sandwich complex consisted of two main parts – one is a peptide bound to a fluorescent protein, and another peptide is immobilized on a cellulose membrane by a cellulose binding domain.
  
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===Source===
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===Design notes===
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The part comes from Clostridium thermocellum genomix sequence.
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The design of proteins attached to the cellulose-binding domain is relatively easy. One of the things that are important when designing a part with this domain is a linker sequence between the domain and the corresponding protein or another biomolecule.
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The sequence of this specific cellulose binding domain came from an article by Yang et al., 2021 [1]. The authors of the article tested two cellulose binding domains and found out, that this domain can successfully be used for paper-based lateral flow immunoassays.
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===Biology===
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Many bacterial and fungal enzymes that hydrolyse insoluble carbohydrates share a familiar structure composed of a catalytic domain linked to carbohydrate-binding module (CBM). Carbohydrate-binding modules (CBMs) are non-catalytic domains that anchor glycoside hydrolases into complex carbohydrates. Clostridium thermocellum produces a multi-enzyme complex of cellulases and hemicellulases, termed the cellulosome, which is organized by the scaffoldin protein CipA.
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C. thermocellum cellulosome. C. thermocellum scaffoldin (CipA) contains nine type I cohesins and thus organizes a multienzyme complex that incorporates nine enzymes. The C-terminal type II dockerin of CipA binds specifically to type II cohesin modules found in cell surface proteins. Individ- ual enzymes may also adhere directly to the bacterium cell envelope by binding the single type I cohesins found in OlpA and OlpC. [[Image:cbd.jpg|400px|thumb|right|Figure 1: C. thermocellum cellulosome. C. thermocellum scaffoldin (CipA) contains nine type I cohesins and thus organizes a multienzyme complex that incorporates nine enzymes. The C-terminal type II dockerin of CipA binds specifically to type II cohesin modules found in cell surface proteins. Individ- ual enzymes may also adhere directly to the bacterium cell envelope by binding the single type I cohesins found in OlpA and OlpC.]]
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https://www.researchgate.net/figure/Structure-of-the-novel-type-I-Coh-Doc-complexes-CohOlpA-Doc918-and-CohOlpC-Doc124A-The_fig2_232766086
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Binding of the cellulosome to the plant cell wall results from the action of CipA family 3 CBM (CBM3), which presents a high affinity for crystalline cellulose.  CBMs that are specific for insoluble cellulose (cellulose binding domain – CBD) represent the predominant category. The CBMs can be grouped into distinctive families on the basis of amino acid sequence similarities. CBM3 is a family of protein modules specific for Gram-positive bacterial families. The proteins comprise of around 150 amino acids. The family of proteins is divided into four subgroups: CBM3a, CBM3b, CBM3c, CBM3d. The major ligand recognised by CBM3as and CBM3bs is crystalline cellulose with an affinity (Kd) of 0,4 uM determined by depletion isotherms.  The family 3a (scaffoldin) and 3b (mainly free enzymes) are closely similar in their primary structures and both types bind strongly to crystalline cellulose [2,3]. Members of the family IIIc, fails to bind crystalline cellulose, but serves in a 'helper' capacity by feeding a single incoming cellulose chain into the active site of the neighbouring catalytic module pending hydrolysis [4,5].
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[[Image:cbd.jpg|400px|thumb|right|Figure 1: C. thermocellum cellulosome. C. thermocellum scaffoldin (CipA) contains nine type I cohesins and thus organizes a multienzyme complex that incorporates nine enzymes. The C-terminal type II dockerin of CipA binds specifically to type II cohesin modules found in cell surface proteins. Individ- ual enzymes may also adhere directly to the bacterium cell envelope by binding the single type I cohesins found in OlpA and OlpC.]]
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===References===
 
===References===
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[1] Yang, J. M., Kim, K. R., Jeon, S., Cha, H. J., & Kim, C. S. (2021). A sensitive paper-based lateral flow immunoassay platform using engineered cellulose-binding protein linker fused with antibody-binding domains. Sensors and Actuators B: Chemical, 329, 129099. https://doi.org/10.1016/j.snb.2020.129099

Latest revision as of 16:59, 13 October 2022


Cellulose Binding domain (CBD)


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design

Vilnius-Lithuania Igem's 2022 team used this part as a novel way for peptide immobilization. The team was working to create an easily accessible nanoplastic detection tool, using peptides, whose interaction with nanoplastic particles would lead to an easily interpretable response. The system itself focused on smaller protein molecules, peptides, which are modified to acquire the ability to connect to the surface of synthetic polymers – plastics. The detection system works when peptides and nanoplastic particles combine and form a "sandwich" complex - one nanoplastic particle is surrounded by two peptides, attached to their respective protein. The sandwich complex consisted of two main parts – one is a peptide bound to a fluorescent protein, and another peptide is immobilized on a cellulose membrane by a cellulose binding domain.

Design notes

The design of proteins attached to the cellulose-binding domain is relatively easy. One of the things that are important when designing a part with this domain is a linker sequence between the domain and the corresponding protein or another biomolecule. The sequence of this specific cellulose binding domain came from an article by Yang et al., 2021 [1]. The authors of the article tested two cellulose binding domains and found out, that this domain can successfully be used for paper-based lateral flow immunoassays.

References

[1] Yang, J. M., Kim, K. R., Jeon, S., Cha, H. J., & Kim, C. S. (2021). A sensitive paper-based lateral flow immunoassay platform using engineered cellulose-binding protein linker fused with antibody-binding domains. Sensors and Actuators B: Chemical, 329, 129099. https://doi.org/10.1016/j.snb.2020.129099