Part:BBa_K3979005
T. aestivum Wild Type Chitinase
Chitinase II precursor derived from Triticum aestivum cultivar Sumai 3. The molecular size and weight of the protein are 24.73 kDa and 719 base pairs.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NotI site found at 712
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 7
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 73
Illegal NgoMIV site found at 621 - 1000COMPATIBLE WITH RFC[1000]
Overview
Chitinases are glycosyl hydrolases (GH) whose catalytic mechanism involves the hydrolysis of the β-1-4-linkage in the N-acetyl-D-glucosamine polymer of chitin, which is a major structural component of fungi. Chitinases are classified into two types based on their cleavage and hydrolysis mechanisms: endochitinase and exo-chitinase. Plants lack chitin, and it has been proposed that plant chitinases play a role in defense response by degrading chitin in invading pathogens’ cell walls. Chitinases isolated from plants have been shown to inhibit the growth of chitin-containing fungi in-vitro and in-vivo, and over-expression of chitinases in plants confers resistance against a variety of fungal pathogens. Chitinases are classified into seven classes (Class I–VII) and contain catalytic domains that define the two major GH families (GH18 and GH19). GH18 chitinases (class III and V) are found in a wide range of organisms, while GH19 chitinases (class I, II, IV, VI, VII) are found mainly in higher plants and are responsible for the majority of chitinolytic activity[1].Chitinase genes code for enzymes that degrade chitin polysaccharides from their reducing end. Plants are significant sources of chitinase proteins where they use these enzymes to degrade chitin for nutrition. Wheat chitinase belongs to an acidic type of class VII chitinase (glycosyl hydrolase family 19), with 77 percent identity to other wheat chitinases of class IV and low-level identity to other plant chitinases, according to molecular phylogenetic investigations [1]. In the present study, the wheat chitinase gene was subcloned and overexpressed in Escherichia coli BL-21 (DE3)[2].
Primer Sequences Used
Protein Structure from RaptorX
The annotated sequence was input into the RaptorX server to give us the predicted 3D structure in the form of a PDB file.
File:Wheat.mp4
Autodock Results
The receptor, here, is our engineered chimeric chitinase and the ligand is the Chitin octamer(CID 24978517).The threshold binding energy is -6kcal/mol which is generally accepted as the cut-off in ligand-binding /docking studies, any value more negative than this is considered significant. So, this protein will show binding with the chitin polymer. The protein structures were prepared before docking by removing water molecules, adding polar hydrogen atoms, and adding Kollman charges. A grid box was created so as to eliminate any surface binding and provide us with better and more reliable results. These modifications are necessary for the efficient binding of the ligand to the protein through non-covalent interactions.
Mode |
Affinity (kcal/mol) |
Dist from RMSD L.B. |
Best mode RMSD U.B. |
1 |
-10.6 |
0 |
0 |
2 |
-10.5 |
6.467 |
11.261 |
3 |
-10.4 |
8.081 |
12.652 |
4 |
-10.0 |
38.776 |
42.149 |
5 |
-9.9 |
6.732 |
11.615 |
6 |
-9.9 |
1.308 |
1.915 |
7 |
-9.7 |
6.283 |
11.248 |
8 |
-9.7 |
5.259 |
9.839 |
9 |
-9.7 |
5.139 |
14.889 |
==References==
- Singh A, Kirubakaran SI, Sakthivel N. Heterologous expression of new antifungal chitinase from wheat. Protein Expr Purif. 2007 Nov;56(1):100-9. doi: 10.1016/j.pep.2007.06.013. Epub 2007 Jul 12. PMID: 17697785.
- Bartholomew ES, Black K, Feng Z, et al. Comprehensive Analysis of the Chitinase Gene Family in Cucumber (Cucumis sativus L.): From Gene Identification and Evolution to Expression in Response to Fusarium oxysporum. Int J Mol Sci. 2019;20(21):5309. Published 2019 Oct 25. doi:10.3390/ijms20215309
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