Difference between revisions of "Part:BBa K1319003"

(→‎Usage and Biology)
(→‎MIT MAHE 2020)
 
(2 intermediate revisions by 2 users not shown)
Line 13: Line 13:
 
===Usage and Biology===
 
===Usage and Biology===
  
 +
K1319003 was used to create [https://parts.igem.org/Part:BBa_K1319020 K1319020], a Galectin-mRFP fusion protein with a C-terminal His tag in the [http://2014.igem.org/Team:Heidelberg/Team/Collaborations Heidelberger expression vector] pSBX1A3.
 +
 +
We also cloned our K1319003 into the pET17 expression vector and expressed all combinations of fusion proteins in E. coli BL21(DE3). An SDS-PAGE showed that all fusion proteins were fully translated:
 
<center>
 
<center>
 
<div class="figure" style="float:{{{align|center}}; margin: 0px 10px 10px 0px; border:{{{border|0px solid #aaa}}};width:{{{width|800px}}};padding:10px 10px 0px 0px;">
 
<div class="figure" style="float:{{{align|center}}; margin: 0px 10px 10px 0px; border:{{{border|0px solid #aaa}}};width:{{{width|800px}}};padding:10px 10px 0px 0px;">
Line 42: Line 45:
 
<partinfo>BBa_K1319003 parameters</partinfo>
 
<partinfo>BBa_K1319003 parameters</partinfo>
 
<!-- -->
 
<!-- -->
 +
 +
==MIT_MAHE 2020==
 +
'''Summary'''
 +
 +
Galectins are proteins of the lectin family, which posess carbonhydrate recognition domains binding specifically to β-galactoside sugar residues. It is encoded by a single gene, LGALS3. It has many physiological functions, such as cell adhesion, cell growth and differentiation, and contributes to the development of cancer, inflammation, fibrosis and others. In the nucleus it acts as a pre-mRNA splicing factor. Together with TRIM16, coordinates the recognition of membrane damage with mobilization of the core autophagy regulators ATG16L1 and BECN1 in response to damaged endomembranes.
 +
 +
==References==
 +
 +
1. Henderson, N. C., & Sethi, T. (2009). The regulation of inflammation by galectin-3. Immunological reviews, 230(1), 160–171. https://doi.org/10.1111/j.1600-065X.2009.00794.x 
 +
 +
2. Haudek, K. C., Spronk, K. J., Voss, P. G., Patterson, R. J., Wang, J. L., & Arnoys, E. J. (2010). Dynamics of galectin-3 in the nucleus and cytoplasm. Biochimica et biophysica acta, 1800(2), 181–189. https://doi.org/10.1016/j.bbagen.2009.07.005
 +
 +
3. Fukushi, J., Makagiansar, I. T., & Stallcup, W. B. (2004). NG2 proteoglycan promotes endothelial cell motility and angiogenesis via engagement of galectin-3 and alpha3beta1 integrin. Molecular biology of the cell, 15(8), 3580–3590. https://doi.org/10.1091/mbc.e04-03-0236

Latest revision as of 18:00, 23 October 2020

human galectin-3, codon optimized for E. coli

Galectin-3 is a 26 kDa protein that binds certain LPS patterns. It especially bind the O-section of the LPS.

Galectins are proteins of the lectin family, which posess carbonhydrate recognition domains binding specifically to β-galactoside sugar residues. In humans, 10 different galectines have been identified, among which is galectin-3.

Galectin-3 has a size of about 31 kDA and is encoded by a single gene, LGALS3. It has many physiological functions, such as cell adhesion, cell growth and differentiation, and contributes to the development of cancer, inflammation, fibrosis and others.

Human galectin-3 is a protein of the lectin-family that was shown to bind the LPS of multiple human pathogens. Some of them, including Pseudomonas aeruginosa protect themselves against the human immune system by mimicking the lipopolysaccharides (LPS) present on human erythrocytes.

Usage and Biology

K1319003 was used to create K1319020, a Galectin-mRFP fusion protein with a C-terminal His tag in the [http://2014.igem.org/Team:Heidelberg/Team/Collaborations Heidelberger expression vector] pSBX1A3.

We also cloned our K1319003 into the pET17 expression vector and expressed all combinations of fusion proteins in E. coli BL21(DE3). An SDS-PAGE showed that all fusion proteins were fully translated:

Pellets of different fusion protein expressions
Expression in the pET17 vector was much more leaky than the expression in the pSBX vectors.


SDS-PAGE of K1319020 expression
The fusion protein was fully translated to the correct molecular mass of 74 kDa.


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 112
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


MIT_MAHE 2020

Summary

Galectins are proteins of the lectin family, which posess carbonhydrate recognition domains binding specifically to β-galactoside sugar residues. It is encoded by a single gene, LGALS3. It has many physiological functions, such as cell adhesion, cell growth and differentiation, and contributes to the development of cancer, inflammation, fibrosis and others. In the nucleus it acts as a pre-mRNA splicing factor. Together with TRIM16, coordinates the recognition of membrane damage with mobilization of the core autophagy regulators ATG16L1 and BECN1 in response to damaged endomembranes.

References

1. Henderson, N. C., & Sethi, T. (2009). The regulation of inflammation by galectin-3. Immunological reviews, 230(1), 160–171. https://doi.org/10.1111/j.1600-065X.2009.00794.x

2. Haudek, K. C., Spronk, K. J., Voss, P. G., Patterson, R. J., Wang, J. L., & Arnoys, E. J. (2010). Dynamics of galectin-3 in the nucleus and cytoplasm. Biochimica et biophysica acta, 1800(2), 181–189. https://doi.org/10.1016/j.bbagen.2009.07.005

3. Fukushi, J., Makagiansar, I. T., & Stallcup, W. B. (2004). NG2 proteoglycan promotes endothelial cell motility and angiogenesis via engagement of galectin-3 and alpha3beta1 integrin. Molecular biology of the cell, 15(8), 3580–3590. https://doi.org/10.1091/mbc.e04-03-0236