Difference between revisions of "Part:BBa K2976004"
Line 2: Line 2:
 
<partinfo>BBa_K2976004 short</partinfo>
 
<partinfo>BBa_K2976004 short</partinfo>
  
−
Granulysin is a member of the saposin-like protein (SAPLIP) family and primarily secreted by cytotoxic T lymphocytes (CTL) and natural killer (NK) cells, which possesses antimicrobial activity against M. tuberculosis and other organisms.
+
Granulysin is a member of the saposin-like protein (SAPLIP) family and primarily secreted by cytotoxic T lymphocytes (CTL) and natural killer (NK) cells, which possesses antimicrobial activity against M. tuberculosis (Mtb) and other organisms [1].
  
−
===Usage===
+
===Usage and Biology===
−
<p>In 2019 CPU_CHINA project, we choose granulysin as a weapon to directly attack extracellular Mtb in blood circulation due to its excellent anti-TB activity.</p>
+
<p>
−
 
+
It is generally recognized that cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells play vital roles in host defense against pathogenic bacteria, which could exhibit cytolytic activity against microbes via producing various antimicrobial substances, such as perforin, granzyme and granulysin [2]. Granulysin is a cationic protein possessing broad antimicrobial activity, especially fatal to Mtb. It directly kills extracellular Mtb by damaging membrane integrity and disrupting lipid metabolism of the bacillus [3]. Moreover, granulysin significantly decreased the number of bacteria in Mtb H37Rv-infected mice model and there was no obvious cytotoxicity of granulysin [4]. Therefore, in 2019 CPU_CHINA project, we choose granulysin as a weapon to directly attack extracellular Mtb in blood circulation due to its excellent anti-TB activity.
−
===Biology===
+
−
<p>It is generally recognized that cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells play vital roles in host defense against pathogenic bacteria, which could exhibit cytolytic activity against microbes via producing various antimicrobial substances, such as perforin, granzyme and granulysin. Granulysin is a cationic protein possessing broad antimicrobial activity, especially fatal to Mtb. It directly kills extracellular Mtb by damaging membrane integrity and disrupting lipid metabolism of the bacillus. Besides, it was reported that there were no obvious changes in proliferation of the granulysin-treated macrophages, which proves that granulysin is non-toxic to human cells.
+
 
</p>
 
</p>
  
 +
===Characterization===
 +
<p>2019 CPU_CHINA attempted to develop a novel strategy to treat tuberculosis based on immune-like cells. Since granulysin is a major part of our project, we conducted research on it and obtained valuable results.</p>
 +
 +
<p>Plasmid (TLR2-P2A-TLR1-T2A-CD14) and plasmid (NF-κB induced promoter-granulysin) were co-transfected into HEK293T cells. Then, we utilized Pam3Cys-Ser-(Lys)4, TLR1/TLR2 agonist to stimulate Toll like receptor (TLR) and its downstream signaling pathway. In our project, the activation of NF-κB can eventually leads to the expression of granulysin. Since the granulysin contains signal peptides, it is eventually destined to be secreted out of cells. Therefore, we determine the concentration of granulysin by ELISA assay. As shown in Figure 1, the concentration of granulysin in the cell culture medium is significantly higher than other groups at 24h (A), 36h (B) and 48h (C).</p>
  
 
<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here

Revision as of 15:58, 20 October 2019

Granulysin

Granulysin is a member of the saposin-like protein (SAPLIP) family and primarily secreted by cytotoxic T lymphocytes (CTL) and natural killer (NK) cells, which possesses antimicrobial activity against M. tuberculosis (Mtb) and other organisms [1].

Usage and Biology

It is generally recognized that cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells play vital roles in host defense against pathogenic bacteria, which could exhibit cytolytic activity against microbes via producing various antimicrobial substances, such as perforin, granzyme and granulysin [2]. Granulysin is a cationic protein possessing broad antimicrobial activity, especially fatal to Mtb. It directly kills extracellular Mtb by damaging membrane integrity and disrupting lipid metabolism of the bacillus [3]. Moreover, granulysin significantly decreased the number of bacteria in Mtb H37Rv-infected mice model and there was no obvious cytotoxicity of granulysin [4]. Therefore, in 2019 CPU_CHINA project, we choose granulysin as a weapon to directly attack extracellular Mtb in blood circulation due to its excellent anti-TB activity.

Characterization

2019 CPU_CHINA attempted to develop a novel strategy to treat tuberculosis based on immune-like cells. Since granulysin is a major part of our project, we conducted research on it and obtained valuable results.

Plasmid (TLR2-P2A-TLR1-T2A-CD14) and plasmid (NF-κB induced promoter-granulysin) were co-transfected into HEK293T cells. Then, we utilized Pam3Cys-Ser-(Lys)4, TLR1/TLR2 agonist to stimulate Toll like receptor (TLR) and its downstream signaling pathway. In our project, the activation of NF-κB can eventually leads to the expression of granulysin. Since the granulysin contains signal peptides, it is eventually destined to be secreted out of cells. Therefore, we determine the concentration of granulysin by ELISA assay. As shown in Figure 1, the concentration of granulysin in the cell culture medium is significantly higher than other groups at 24h (A), 36h (B) and 48h (C).

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 470
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]