title: Swapping the chitin-binding domain in Bacillus chitinases improves the substrate binding affinity and conformational stability.
abstract: Chitinase from Bacillus thuringiensis and Bacillus licheniformis consisting of an N-terminal catalytic domain (GH18) and a C-terminal chitin-binding domain (ChBD), were cloned and characterised. In order to study the importance of individual domains, chimeric chitinases (BtGH-BliChBD and BliGH-BtChBD) were constructed using domain swapping as a strategy to exchange the CBD of BtGH-ChBD with that of BliGH-ChBD and vice versa. Both chimeric chitinases showed increased affinity to colloidal chitin. BtGH-BliChBD was different from the three other chitinases studied concerning optimum temperature and pH. Additionally, BtGH-BliChBD and BliGH-BtChBD showed significant improvement in functional stability, conformational stability, and binding ability towards insoluble chitinous substrates compared to those of the native chitinases.
authors.affiliation: Department of Plant Sciences, University of Hyderabad, Hyderabad, India.
authors.forename: Chilukoti
authors.initials: C
authors.lastname: Neeraja
authors.forename: Rajagopal
authors.initials: R
authors.lastname: Subramanyam
authors.forename: Bruno M
authors.initials: BM
authors.lastname: Moerschbacher
authors.forename: Appa Rao
authors.initials: AR
authors.lastname: Podile
mesh_terms: D001407:Bacillus / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D002686:Chitin / Q000378:metabolism*; D002688:Chitinases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D004795:Enzyme Stability; D006868:Hydrolysis; D016297:Mutagenesis, Site-Directed / Q000379:methods; D011485:Protein Binding; D011487:Protein Conformation; D015202:Protein Engineering; D054730:Protein Interaction Domains and Motifs / Q000235:genetics / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000138:chemical synthesis / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013379:Substrate Specificity; D013816:Thermodynamics
chemical_list: D011993:Recombinant Fusion Proteins; D002686:Chitin; D002688:Chitinases