Difference between revisions of "Part:BBa K5185008"

 
Line 3: Line 3:
 
<partinfo>BBa_K5185008 short</partinfo>
 
<partinfo>BBa_K5185008 short</partinfo>
  
−
CBMxx is yet another family of CBM that binds alginate. Alginate possesses high biocompatibility, low toxicity, and could provide a moist wound environment and promote wound healing when delivered in a hydrogel wound dressing formation
+
Carbohydrate-Binding Modules from Vibrio breoganii (VbCBMxx) are specialized domains found in enzymes that degrade marine polysaccharides. These CBMs enhance enzyme binding to substrates like agar, laminarin, and alginate, improving degradation efficiency. By incorporating VbCBMxx into proteins, we can target marine biomass for conversion or modification and may be applicable in marine biotechnology and bioprocessing.
 +
 
 +
Carbohydrate-binding modules (CBMs) are modules with specificity toward different types of carbohydrates when assisting in carbohydrate degradation. Other than VbCBMxx, this part collection includes other CBMs such as CBM2, CBM3, CBM5, and human integrin domains such as α1 and α2.
 +
 
 +
This is part of a part collection of CBMs aimed to provide first aid wound dressings with enhanced antimicrobial functions and a wider and more complex application, where we characterize bacterial cellulose modification methods and constructs using CBMs as a binding domain, linking HNPs such as HNP1 and HNP4 to carbohydrates such as cellulose and chitosan. By fusing VbCBMxx to functional proteins, sodium-alginate-containing materials can be effective targets for modification or degradation. The part collection includes: Cellulose binding matrices CBM2 (<partinfo>BBa_K4011001 </partinfo>) which binds to trehalose, CBM3 (<partinfo>BBa_K4011000</partinfo>) which binds to fibrin, CBM5 (<partinfo>BBa_K5185002</partinfo>) which binds to chitosan, and VbCBMxx (<partinfo>BBa_K5185008</partinfo>) which binds to sodium alginate. Human integrins α1 domain (<partinfo>BBa_K5185003 </partinfo>) and α2 domain (<partinfo>BBa_K5185004</partinfo>), linking functional proteins to collagen. This part collection can help to achieve modification of cellulose membranes using different modification/functionalization proteins.
 +
 
 +
Since sodium alginate is cheaper than its other wound dressing counterparts and it does not present secondary damage to wounds, it could be used for wound dressings on wounds that prioritize affordability and wound protection.
  
−
<!-- Add more about the biology of this part here
 
 
===Usage and Biology===
 
===Usage and Biology===
 +
In its natural habitat, Vibrio breoganii produces enzymes with VbCBMxx domains to degrade polysaccharides found in algae and seaweeds. VbCBMxx binds specifically to marine carbohydrates, allowing the bacterium to utilize these polysaccharides as nutrient sources. VbCBMxx also exhibits the desired specificity to alginate. The recent discovery of VbCBMxx by Mei et al. expanded the toolbox of alginate-binding proteins.
 +
 +
===Source===
 +
VbCBMxx modules are derived from the marine bacterium Vibrio breoganii.
  
−
<!-- -->
+
===Results===
−
<span class='h3bb'>Sequence and Features</span>
+
We obtained a fusion protein composed of a binding domain protein linked to a fluorescent protein, naming it VbCBMxx-gfasPurple.
−
<partinfo>BBa_K5185008 SequenceAndFeatures</partinfo>
+
Figure A shows the results of the SDS-PAGE analysis of the target fusion proteins (binding domains linked to chromoproteins). The VbCBMxx-gfasPurple fusion protein was expressed in E. coli SHuffle T7. VbCBMxx-gfasPurple has a molecular weight of 49.5 kDa and is successfully expressed.
 +
The color of VbCBMxx-gfasPurple suspended in 20mM Tris-HCl under sunlight is shown in Figure B, expressing the correct color purple.
 +
VbCBMxx's ability to bind to sodium alginate is assessed, as shown in Figure C. The results indicate that VbCBMxx can successfully bind to sodium alginate material, as shown by the deeper purple color observed under sunlight.
  
 +
<html>
 +
<img style="display: block;
 +
    width: 60%;height: 60%;" src="https://static.igem.wiki/teams/5185/part-org/binding-domain-sodium-alginate.png" text-align="center"><div>Figure 1: </div></html>
 +
<br>(a) SDS-PAGE identification of BDC shows the results of SDS-PAGE analysis of the target fusion proteins(BDC). All fusion proteins were expressed in E. coli BL21 (DE3), except for CBMxx-gfasPurple, which was expressed in the SHuffle T7 strain due to its low solubility in BL21 (DE3). α1-mRFP, α2-eforRed, CBM2-mTurquoise, CBM3-sfGFP, CBM5-fwYellow, and CBMxx-gfasPurple each have molecular weights of 49.2 kDa, 49.1 kDa, 40 kDa, 46kDa, 32.l kDa, and 49.5 kDa, respectively.
 +
<br>(b)Yellow-light detection of BDC shows the color of each fusion protein suspended in 20 mM Tris HCl under bluelight.
 +
<br>(c) Binding efficiencies of cellulose binding domains CBM2 and CBM3 toward cellulose gauze; binding efficiencies of collagen binding domains α1 and α2, chitosan binding domain CBM5, and alginate binding domain VbCBMxx toward respective hydrogel materials.
  
−
<!-- Uncomment this to enable Functional Parameter display
+
===Reference===
−
===Functional Parameters===
+
<br> Armenta, S., Moreno-Mendieta, S., Sánchez-Cuapio, Z., Sánchez, S., & Rodríguez-Sanoja, R. (2017). Advances in molecular engineering of carbohydrate-binding modules. Proteins, 85(9), 1602–1617.
−
<partinfo>BBa_K5185008 parameters</partinfo>
+
<br> Han, Y., Gao, P., Yu, W., & Lu, X. (2017). Thermostability enhancement of chitosanase CsnA by fusion a family 5 carbohydrate-binding module. Biotechnology letters, 39(12), 1895–1901. https://doi.org/10.1007/s10529-017-2406-2
−
<!-- -->
+
<br> Heino, J., Siljamäki, E. (2023). Integrins α1β1 and α2β1: The Generalist Collagen Receptors. In: Gullberg, D., Eble, J.A. (eds) Integrins in Health and Disease. Biology of Extracellular Matrix, vol 13. Springer, Cham. https://doi.org/10.1007/978-3-031-23781-2_1
 +
<br> Mei, X., Tao, W., Sun, H., Liu, G., Chen, G., Zhang, Y., Xue, C., & Chang, Y. (2024). Characterization and structural identification of a novel alginate-specific carbohydrate-binding module (CBM): The founding member of a new CBM family. International journal of biological macromolecules, 277(Pt 3), 134221. https://doi.org/10.1016/j.ijbiomac.2024.134221

Revision as of 19:45, 1 October 2024


Carbohydrate -binding modules (CBMxx)

Carbohydrate-Binding Modules from Vibrio breoganii (VbCBMxx) are specialized domains found in enzymes that degrade marine polysaccharides. These CBMs enhance enzyme binding to substrates like agar, laminarin, and alginate, improving degradation efficiency. By incorporating VbCBMxx into proteins, we can target marine biomass for conversion or modification and may be applicable in marine biotechnology and bioprocessing.

Carbohydrate-binding modules (CBMs) are modules with specificity toward different types of carbohydrates when assisting in carbohydrate degradation. Other than VbCBMxx, this part collection includes other CBMs such as CBM2, CBM3, CBM5, and human integrin domains such as α1 and α2.

This is part of a part collection of CBMs aimed to provide first aid wound dressings with enhanced antimicrobial functions and a wider and more complex application, where we characterize bacterial cellulose modification methods and constructs using CBMs as a binding domain, linking HNPs such as HNP1 and HNP4 to carbohydrates such as cellulose and chitosan. By fusing VbCBMxx to functional proteins, sodium-alginate-containing materials can be effective targets for modification or degradation. The part collection includes: Cellulose binding matrices CBM2 (BBa_K4011001) which binds to trehalose, CBM3 (BBa_K4011000) which binds to fibrin, CBM5 (BBa_K5185002) which binds to chitosan, and VbCBMxx (BBa_K5185008) which binds to sodium alginate. Human integrins α1 domain (BBa_K5185003) and α2 domain (BBa_K5185004), linking functional proteins to collagen. This part collection can help to achieve modification of cellulose membranes using different modification/functionalization proteins.

Since sodium alginate is cheaper than its other wound dressing counterparts and it does not present secondary damage to wounds, it could be used for wound dressings on wounds that prioritize affordability and wound protection.

Usage and Biology

In its natural habitat, Vibrio breoganii produces enzymes with VbCBMxx domains to degrade polysaccharides found in algae and seaweeds. VbCBMxx binds specifically to marine carbohydrates, allowing the bacterium to utilize these polysaccharides as nutrient sources. VbCBMxx also exhibits the desired specificity to alginate. The recent discovery of VbCBMxx by Mei et al. expanded the toolbox of alginate-binding proteins.

Source

VbCBMxx modules are derived from the marine bacterium Vibrio breoganii.

Results

We obtained a fusion protein composed of a binding domain protein linked to a fluorescent protein, naming it VbCBMxx-gfasPurple. Figure A shows the results of the SDS-PAGE analysis of the target fusion proteins (binding domains linked to chromoproteins). The VbCBMxx-gfasPurple fusion protein was expressed in E. coli SHuffle T7. VbCBMxx-gfasPurple has a molecular weight of 49.5 kDa and is successfully expressed. The color of VbCBMxx-gfasPurple suspended in 20mM Tris-HCl under sunlight is shown in Figure B, expressing the correct color purple. VbCBMxx's ability to bind to sodium alginate is assessed, as shown in Figure C. The results indicate that VbCBMxx can successfully bind to sodium alginate material, as shown by the deeper purple color observed under sunlight.

Figure 1:

(a) SDS-PAGE identification of BDC shows the results of SDS-PAGE analysis of the target fusion proteins(BDC). All fusion proteins were expressed in E. coli BL21 (DE3), except for CBMxx-gfasPurple, which was expressed in the SHuffle T7 strain due to its low solubility in BL21 (DE3). α1-mRFP, α2-eforRed, CBM2-mTurquoise, CBM3-sfGFP, CBM5-fwYellow, and CBMxx-gfasPurple each have molecular weights of 49.2 kDa, 49.1 kDa, 40 kDa, 46kDa, 32.l kDa, and 49.5 kDa, respectively.
(b)Yellow-light detection of BDC shows the color of each fusion protein suspended in 20 mM Tris HCl under bluelight.
(c) Binding efficiencies of cellulose binding domains CBM2 and CBM3 toward cellulose gauze; binding efficiencies of collagen binding domains α1 and α2, chitosan binding domain CBM5, and alginate binding domain VbCBMxx toward respective hydrogel materials.

Reference


Armenta, S., Moreno-Mendieta, S., Sánchez-Cuapio, Z., Sánchez, S., & Rodríguez-Sanoja, R. (2017). Advances in molecular engineering of carbohydrate-binding modules. Proteins, 85(9), 1602–1617.
Han, Y., Gao, P., Yu, W., & Lu, X. (2017). Thermostability enhancement of chitosanase CsnA by fusion a family 5 carbohydrate-binding module. Biotechnology letters, 39(12), 1895–1901. https://doi.org/10.1007/s10529-017-2406-2
Heino, J., Siljamäki, E. (2023). Integrins α1β1 and α2β1: The Generalist Collagen Receptors. In: Gullberg, D., Eble, J.A. (eds) Integrins in Health and Disease. Biology of Extracellular Matrix, vol 13. Springer, Cham. https://doi.org/10.1007/978-3-031-23781-2_1
Mei, X., Tao, W., Sun, H., Liu, G., Chen, G., Zhang, Y., Xue, C., & Chang, Y. (2024). Characterization and structural identification of a novel alginate-specific carbohydrate-binding module (CBM): The founding member of a new CBM family. International journal of biological macromolecules, 277(Pt 3), 134221. https://doi.org/10.1016/j.ijbiomac.2024.134221