Domain architecture divergence leads to functional divergence in binding and catalytic domains of bacterial and fungal cellobiohydrolases
| Autor: | Ryota Iino, Taku Uchiyama, Akihiko Nakamura, Kenji Mizutani, Akasit Visootsat, Satoshi Kaneko, Kiyohiko Igarashi, Daiki Ishiwata, Takeshi Murata |
|---|---|
| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
Models Molecular microscopic imaging Protein Conformation carbohydrate-binding protein single-molecule biophysics Trichoderma reesei Cellulase Cellobiose Crystallography X-Ray Biochemistry Substrate Specificity Fungal Proteins 03 medical and health sciences chemistry.chemical_compound glycoside hydrolase family 6 Bacterial Proteins Protein Domains Catalytic Domain Cellulose 1 4-beta-Cellobiosidase Glycoside hydrolase glycoside hydrolase Cellulose Molecular Biology Cellulomonas cellulase Cellulomonas fimi Binding Sites 030102 biochemistry & molecular biology biology single-molecule observation Chemistry Cell Biology Processivity processivity biology.organism_classification molecular imaging 030104 developmental biology Hypocreales biology.protein Biophysics Enzymology Linker Functional divergence Protein Binding |
| Zdroj: | The Journal of Biological Chemistry |
| ISSN: | 1083-351X |
| Popis: | Cellobiohydrolases directly convert crystalline cellulose into cellobiose and are of biotechnological interest to achieve efficient biomass utilization. As a result, much research in the field has focused on identifying cellobiohydrolases that are very fast. Cellobiohydrolase A from the bacterium Cellulomonas fimi (CfCel6B) and cellobiohydrolase II from the fungus Trichoderma reesei (TrCel6A) have similar catalytic domains (CDs) and show similar hydrolytic activity. However, TrCel6A and CfCel6B have different cellulose-binding domains (CBDs) and linkers: TrCel6A has a glycosylated peptide linker, whereas CfCel6B's linker consists of three fibronectin type 3 domains. We previously found that TrCel6A's linker plays an important role in increasing the binding rate constant to crystalline cellulose. However, it was not clear whether CfCel6B's linker has similar function. Here we analyze kinetic parameters of CfCel6B using single-molecule fluorescence imaging to compare CfCel6B and TrCel6A. We find that CBD is important for initial binding of CfCel6B, but the contribution of the linker to the binding rate constant or to the dissociation rate constant is minor. The crystal structure of the CfCel6B CD showed longer loops at the entrance and exit of the substrate-binding tunnel compared with TrCel6A CD, which results in higher processivity. Furthermore, CfCel6B CD showed not only fast surface diffusion but also slow processive movement, which is not observed in TrCel6A CD. Combined with the results of a phylogenetic tree analysis, we propose that bacterial cellobiohydrolases are designed to degrade crystalline cellulose using high-affinity CBD and high-processivity CD. |
| Databáze: | OpenAIRE |
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