Journal
MOLECULAR MICROBIOLOGY
Volume 96, Issue 1, Pages 14-27Publisher
WILEY-BLACKWELL
DOI: 10.1111/mmi.12915
Keywords
-
Categories
Funding
- KAKENHI (Japan Society for the Promotion of Science) [20380175, 24658281]
- RITE (Research Institute of Innovative Technology for the Earth)
- Grants-in-Aid for Scientific Research [20380175, 24658281] Funding Source: KAKEN
Ask authors/readers for more resources
Hydrophobins are amphipathic proteins secreted by filamentous fungi. When the industrial fungus Aspergillus oryzae is grown in a liquid medium containing the polyester polybutylene succinate co-adipate (PBSA), it produces RolA, a hydrophobin, and CutL1, a PBSA-degrading cutinase. Secreted RolA attaches to the surface of the PBSA particles and recruits CutL1, which then condenses on the particles and stimulates the hydrolysis of PBSA. Here, we identified amino acid residues that are required for the RolA-CutL1 interaction by using site-directed mutagenesis. We quantitatively analyzed kinetic profiles of the interactions between RolA variants and CutL1 variants by using a quartz crystal microbalance (QCM). The QCM analyses revealed that Asp142, Asp171 and Glu31, located on the hydrophilic molecular surface of CutL1, and His32 and Lys34, located in the N-terminus of RolA, play crucial roles in the RolA-CutL1 interaction via ionic interactions. RolA immobilized on a QCM electrode strongly interacted with CutL1 (K-D=6.5nM); however, RolA with CutL1 variants, or RolA variants with CutL1, showed markedly larger K-D values, particularly in the interaction between the double variant RolA-H32S/K34S and the triple variant CutL1-E31S/D142S/D171S (K-D=78.0nM). We discuss a molecular prototype model of hydrophobin-based enzyme recruitment at the solid-water interface.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available