Part:BBa_K3979009

ChiB gene of Serratia marcescens of QMB1466

Chitinase B gene of Serratia marcescens of QMB1466. The molecular size and weight of the protein are 55.46438 kDa and 1524 base pairs.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 7
  Illegal XhoI site found at 1519
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Overview

The chitinase enzyme hydrolyzes insoluble chitin to its oligo and monomeric components. Chitinase proteins are abundant in microorganisms such as bacteria, which use these enzymes to degrade chitin for nutrition. Chitinases are classified as endochitinases or exochitinases. Endochitinases cleave chitin at internal sites to produce GlcNAc multimers. Exochitinases catalyze the progressive hydrolysis of chitin to produce GlcNAc, chitobiose, or chitotriose. Chitinases are classified into different glycoside hydrolase (GH) families based on their amino acid sequences, such as GH18, GH19, and GH20. The GH18 family contains the majority of bacterial chitinases. Based on amino acid sequence homology of the individual catalytic domains, bacterial GH18 chitinases are classified into three major subfamilies, A, B, and C. Chitinolytic bacteria are found in a variety of habitats and decompose chitin in both aerobic and anaerobic conditions[2]. Chitin, after cellulose, is the second most abundant natural resource. Chitinases have been suggested to be involved in pathogen-antagonist interaction during the biological control progress of plant pathogenic fungi. Serratia marcescens is one of the most efficient bacteria for chitin breakdown due to a wide range of chitinase and chitin-binding proteins[1].

Primer Sequences Used

Fig. 1. ChiB Forward Primer

Fig. 2. ChiB Reverse Primer

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.

Structure

Autodock Results

The annotated sequence was input into the RaptorX server to give us the predicted 3D structure in the form of a PDB file. 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	-12.2	0	0
2	-11.9	23.618	29.978
3	-11.6	29.247	33.375
4	-11.6	4.887	11.213
5	-11.4	5.934	12.749
6	-11.1	10.591	15.896
7	-11	12.310	18.205
8	-10.9	28.933	34.958
9	-10.9	67.923	73.297