Elucidating Cellulose Film Properties and Enzyme Kinetics Using MP-SPR

Cellulose and its derivatives are widely acknowledged for their versatility, biodegradability, and cost-effectiveness, making them ideal candidates for various biomedical applications, such as wound dressings, drug delivery systems, and surgical sutures. Multi-Parametric Surface Plasmon Resonance (MP-SPR) has been extensively utilized to study the properties of thin films composed of cellulose and its derivatives. By employing a multi-wavelength approach, MP-SPR enables the precise determination of both the thickness and refractive index of these films [1,2]. This method not only allows for the accurate measurement of single or multilayer thickness [1] but also facilitates the determination of hydration of the films [2]. Furthermore, MP-SPR has been instrumental in exploring the interactions between proteins and modified cellulose thin films ​[1].

Recently, MP-SPR was employed to investigate the enzymatic degradation of xylan-modified cellulose by xylanase. In this study, the researchers created thin films on MP-SPR sensor slides, followed by xylan modification, and subsequently measured the thickness of these multilayers​ [3]. Schaubeder et al. used MP-SPR to first calculate the surface coverage of the xylan-modified cellulose on the sensor surface. To determine surface coverage, they employed the de Feijter’s equation [3, 4]​, which has been extensively used to calculate the protein surface coverage on the MP-SPR sensor. The xylan-modified cellulose thin films were then subjected to enzymatic degradation using endo-1,4-β-xylanase. The MP-SPR sensogram indicated a decrease in the thickness of the xylan-cellulose thin film upon xylanase treatment, with higher xylanase concentrations leading to more significant reductions in layer thickness [3]. This method could be further used to study enzyme degradation in situ. Schaubeder et al. focused on elucidating the enzyme-substrate kinetics of a single xylanase interacting with the modified cellulose thin films.

Enzymatic kinetics are influenced by various factors, and traditional solution-based assays often limit the understanding of heterogeneous surface interactions between enzymes and substrates. Investigating enzyme kinetics using a heterogeneous surface reaction approach provides deeper insights into the activity of xylanases, but it requires a different analytical strategy​. Schaubeder et al. extended the study of xylanase and modified cellulose kinetics using MP-SPR [5]. A multilayer of xylan-modified cellulose was coated on an SPR sensor slide to serve as the substrate for studying xylanase kinetics. For kinetic simulations, they applied a combination of Langmuir and Michaelis-Menten concepts. By integrating these models with data obtained from MP-SPR measurements, the authors were able to shed light on the complex dynamics involved in heterogeneous enzymatic surface reactions. Schaubeder et al. proposed a modified model for these heterogeneous enzymatic surface kinetics​ [1]. The authors validated their model using modified cellulose thin films and enzymes, studying the influence of factors such as flow rate and layer thickness on the kinetic parameters [5]. Additionally, the model was validated by degrading non-cellulosic polyhydroxybutyrate (PHB) polymer with a PHB depolymerase.

These works of Schaubeder et al. provide an excellent system for further studying enzyme kinetics by introducing more heterogeneity into the system, enabling the acquisition of kinetic information that more closely resembles native systems. The referenced articles here provide excellent workflow for creating cellulose thin films on the sensor surface, study their interaction with biomolecules, hydration, and degradation using MP-SPR.

References:

1. Sampl et al. 2019. Multilayer Density Analysis of Cellulose Thin Films.
2. Kontturi et al. 2013. Specific water uptake of thin films from nanofibrillar cellulose.
3. Schaubeder et al. 2022. Xylan-cellulose thin film platform for assessing xylanase activity.
4. Solin et al.  2023. Cannabis detection with solid sensors and paper-based immunoassays by conjugating antibodies to nanocellulose.
5. Schaubeder et al. 2024. Deciphering heterogeneous enzymatic surface reactions on xylan using surface plasmon resonance spectroscopy.