Difference between revisions of "Part:BBa K4990010"

(How does it work?)
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HlpA, as suggested by its name "Histone-like protein A", binds to DNA similar to histones. Its overall morphology resembles a crab claw, allowing it to non-specifically and tightly bind to the minor groove of DNA using its ribbon-like β-fold region. This results in significant DNA bending, DNA compaction, and negative supercoiling. S. bovis, through an unknown mechanism, secretes it extracellularly, acting as an anchorless protein. It becomes a target for the humoral immune system during infections. By linking bacterial lipoteichoic acid (LTA) and HSPG on colon epithelial cells, it mediates bacterial adhesion to colon tumor cells.
 
HlpA, as suggested by its name "Histone-like protein A", binds to DNA similar to histones. Its overall morphology resembles a crab claw, allowing it to non-specifically and tightly bind to the minor groove of DNA using its ribbon-like β-fold region. This results in significant DNA bending, DNA compaction, and negative supercoiling. S. bovis, through an unknown mechanism, secretes it extracellularly, acting as an anchorless protein. It becomes a target for the humoral immune system during infections. By linking bacterial lipoteichoic acid (LTA) and HSPG on colon epithelial cells, it mediates bacterial adhesion to colon tumor cells.
  
Wu et al. [6] found that through their research on chitosan-fused rigid linker peptides, that rigid linker peptides exhibit self-cleaving properties under specific conditions. At pH=6-7, cleavage occurs. It's hypothesized that the EAAAK amino acid arrangement can form a stable hydrophilic α-helical structure. The forces at play can be attributed to the Glu-...Lys+ salt bridges between Glu and Lys. Hydrogen bonds are pivotal in forming these salt bridges, so pH greatly influences the stability of the salt bridges. At neutral pH, the strength of salt bridges is at its weakest, making cleavage more likely. Conversely, in high salt environments, the ionic strength can stabilize these bridges, preventing cleavage [7].
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Wu et al. [7] found that through their research on chitosan-fused rigid linker peptides, that rigid linker peptides exhibit self-cleaving properties under specific conditions. At pH=6-7, cleavage occurs. It's hypothesized that the EAAAK amino acid arrangement can form a stable hydrophilic α-helical structure. The forces at play can be attributed to the Glu-...Lys+ salt bridges between Glu and Lys. Hydrogen bonds are pivotal in forming these salt bridges, so pH greatly influences the stability of the salt bridges. At neutral pH, the strength of salt bridges is at its weakest, making cleavage more likely. Conversely, in high salt environments, the ionic strength can stabilize these bridges, preventing cleavage [8].
  
  
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For mammalian cells, whose membranes are mostly neutral, FK-13's interaction with membranes is comparatively weak, and cells are less susceptible to damage. Furthermore, FK-13 can enhance the rigidity of epithelial cell membranes and reduce their permeability, thus minimizing bacterial attacks. Additionally, FK-13 possesses an amphipathic helical structure, which under hydrophobic conditions promotes oligomerization. The potency of the immune response is directly proportional to the α-helical structure of the oligomers. When there are more α-helices present, the antibacterial activity is stronger[8].
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For mammalian cells, whose membranes are mostly neutral, FK-13's interaction with membranes is comparatively weak, and cells are less susceptible to damage. Furthermore, FK-13 can enhance the rigidity of epithelial cell membranes and reduce their permeability, thus minimizing bacterial attacks. Additionally, FK-13 possesses an amphipathic helical structure, which under hydrophobic conditions promotes oligomerization. The potency of the immune response is directly proportional to the α-helical structure of the oligomers. When there are more α-helices present, the antibacterial activity is stronger[9].
 
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===Reference===
 
===Reference===

Revision as of 04:37, 12 October 2023


Tumour-Targeting Peptide

TO KNOW ABOUT IT!

In the wake of advancements in molecular biology, numerous proteins and peptides with active functions have been discovered or invented for pharmaceutical applications. Recombinant technology, characterized by its high expression levels and ease of operation, has been extensively applied in biomedicine and related fields. To obtain multi-target, multifunctional active proteins, it is requisite to link and fuse two or more proteins with known functions. This method of obtaining bifunctional or multifunctional fusion proteins has become one of the new approaches for developing new drugs and researching bioproducts, especially widely used in the preparation of bispecific single-chain variable fragments (scFv) or antibody-drug conjugates[1-5].

Fusion proteins are principally composed of two parts: the functional protein and the linker peptide. The functional protein is the original protein intended for fusion, typically with known structure and function, posing no issues in selection. However, due to the significance of the linker peptide in the overall structure of the fusion protein, its selection and design require thoughtful research to ensure the overall activity of the fusion protein remains unchanged[2]. Consequently, research on linker peptides has gradually come into focus.

Usage in short

You can use it to target and kill CRC.

What is it

This is the structure of (Tumor-Targeting Peptide)TTP, which is consited of HlpA, GPNG ,Linker B, FK-13.

HlpA targets HSPG on CRC, Linker A is a rigid and cleavage linker, FK-13 has antimicrobial activity.

How does it work?

In 2006, Tjalsma employed the Highly accurate tandem MS method to identify a protein named Histone-like protein A (HlpA). This discovery highlighted a bridge between Streptococcus bovis and colorectal cancer. It is postulated that S. bovis establishes a bond with colorectal cancer cell surfaces via heparan sulfate-proteoglycans (HSPG) mediated by HlpA [1].

By 2009, Boleij confirmed that Streptococcus gallolyticus binds to the surface of colorectal cancer cells through HlpA interacting with HSPG [2].

In 2016, O'Neil unveiled the crystal structure of Hlp, revealing its crab-claw-like architecture. The claw section's basic amino acids can bind with DNA and simultaneously with heparin [3].

In 2018, Chun Loong Ho harnessed the binding of HlpA to HSPG to construct an engineered Escherichia coli that specifically targets colorectal cancer, thus marking the commencement of the HlpA targeting system [4].

In 2022, the iGEM22_LZU-CHINA team, utilizing synthetic biology, developed an Escherichia coli that targets colorectal cancer, employing the method of HlpA targeting tumor cell surface HSPG [5].

By 2023, Tang engineered a probiotic using HlpA targeting and Azurin-induced cytotoxicity, demonstrating promising therapeutic efficacy in colorectal cancer mice models [6].


control
Figure A:Crystal structure of HlpA(O'Neil 2016) Figure B:Engineered EcN treat CRC(Chun 2018)
HlpA, as suggested by its name "Histone-like protein A", binds to DNA similar to histones. Its overall morphology resembles a crab claw, allowing it to non-specifically and tightly bind to the minor groove of DNA using its ribbon-like β-fold region. This results in significant DNA bending, DNA compaction, and negative supercoiling. S. bovis, through an unknown mechanism, secretes it extracellularly, acting as an anchorless protein. It becomes a target for the humoral immune system during infections. By linking bacterial lipoteichoic acid (LTA) and HSPG on colon epithelial cells, it mediates bacterial adhesion to colon tumor cells.

Wu et al. [7] found that through their research on chitosan-fused rigid linker peptides, that rigid linker peptides exhibit self-cleaving properties under specific conditions. At pH=6-7, cleavage occurs. It's hypothesized that the EAAAK amino acid arrangement can form a stable hydrophilic α-helical structure. The forces at play can be attributed to the Glu-...Lys+ salt bridges between Glu and Lys. Hydrogen bonds are pivotal in forming these salt bridges, so pH greatly influences the stability of the salt bridges. At neutral pH, the strength of salt bridges is at its weakest, making cleavage more likely. Conversely, in high salt environments, the ionic strength can stabilize these bridges, preventing cleavage [8].


FK-13 exhibits antibacterial effects against both Gram-positive and Gram-negative bacteria. Through electrostatic interactions, the positively charged FK-13 connects with negatively charged bacteria, inserting into bacterial cell membranes and leading to the leakage of their contents, resulting in cell death.

Currently, two hypotheses support the mechanism by which FK-13 disrupts cell membranes: the carpet model, based on FK-13's cationic amino acids interacting with the phospholipid head groups in the cell membrane, which can form a carpet-like structure on the membrane surface, ultimately compromising membrane integrity. Changes in helical structure, charge, and hydrophobicity influence its antibacterial activity. On the other hand, the toroidal pore model suggests that due to membrane surface tension and curvature, FK-13 induces the formation of toroidal pores in bacterial membranes, causing leakage of bacterial contents and leading to bacterial death.

For mammalian cells, whose membranes are mostly neutral, FK-13's interaction with membranes is comparatively weak, and cells are less susceptible to damage. Furthermore, FK-13 can enhance the rigidity of epithelial cell membranes and reduce their permeability, thus minimizing bacterial attacks. Additionally, FK-13 possesses an amphipathic helical structure, which under hydrophobic conditions promotes oligomerization. The potency of the immune response is directly proportional to the α-helical structure of the oligomers. When there are more α-helices present, the antibacterial activity is stronger[9].

Reference

[1] Tao, L., Gao, M., & Zhou, H. (2015). Research progress on novel antibody-chemotherapy drug conjugates. Journal of Pharmaceutical Biotechnology, 22(3), 253-258.

[2] Gustavsson M,Lehtio J,Denman S,et al. Stable linker peptides for a cellulose-binding domain-lipase fusion protein expressed in pichia pastoris[J]. Protein Eng,2001,14(9):711-715.

[3] Wang SH,Zheng CJ,Liu Y,et al. Construction of multiform scFv antibodies using linker peptide[J]. J Genetics and Genomics,2008,35:313-316.

[4] Zhang JH,Yun J,Shang ZG,et al. Design and optimization of a linker for fusion protein construction[J]. Progr Natur Sci,2009,19: 1197-1200.

[5] Shan D,Press OW,Tsu TT,et al. Characterization of scFv-Ig constructs generated from the Anti-CD20 mAb 1F5 using linker peptides of varying lengths[J].J Immunology,2014,162:6589-6595.

[6]Tang, H., Zhou, T., Jin, W., Zong, S., Mamtimin, T., Salama, E. S., ... & Li, X. (2023). Tumor-targeting engineered probiotic Escherichia coli Nissle 1917 inhibits colorectal tumorigenesis and modulates gut microbiota homeostasis in mice. Life Sciences, 324, 121709.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 282
  • 1000
    COMPATIBLE WITH RFC[1000]