Direct observation of poly(3-hydroxybutyrate) depolymerase adsorbed on polyester thin film by atomic force microscopy

Area of Science:

• Biochemistry
• Materials Science
• Microbiology

Background:

• Poly[(R)-3-hydroxybutyrate] (PHB) depolymerases are enzymes capable of degrading PHB.
• Poly(L-lactide) (PLLA) is a biodegradable polyester with various applications.
• Understanding enzyme-substrate interactions is crucial for biodegradation studies.

Purpose of the Study:

• To directly observe the adsorption behavior of PHB depolymerases on a PLLA thin film.
• To investigate the surface morphology changes induced by enzyme adsorption.
• To elucidate the interaction mechanism between PHB depolymerases and PLLA.

Main Methods:

• Preparation of amorphous PLLA thin films (100 nm) on silicon wafers using spin-casting.
• Treatment of PLLA films at 220°C and quenching to achieve an amorphous state.
• Atomic Force Microscopy (AFM) for direct observation of enzyme adsorption and surface topography.
• Enzyme removal using a 40% ethanol aqueous solution.

Main Results:

• PHB depolymerases adsorbed in an elliptical shape on the amorphous PLLA surface.
• A small ridge was observed around each adsorbed enzyme molecule.
• Small hollows were detected on the PLLA film surface after enzyme removal.
• These findings suggest a direct interaction between PHB depolymerases and PLLA molecules.

Conclusions:

• PHB depolymerases interact with PLLA molecules upon adsorption.
• The adsorption process leads to localized surface modifications, forming hollows.
• This study provides direct visual evidence of enzyme-substrate interaction at the nanoscale.