Surface-Selective SHG Detection for Protein Conformation Analysis
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Solution Overview
Problem
Current high throughput screening technologies for drug discovery face limitations, including the need for sophisticated light sources and detectors sensitive to misalignment, photo-bleaching issues, and difficulty in identifying allosteric modulators for 'undruggable' protein targets, which are challenging due to their large contact areas or high affinity ligands.
Innovation Solution
The development of methods and systems using second harmonic generation (SHG) for high throughput analysis of conformational changes in biological entities, involving surface-selective optical techniques with total internal reflection to efficiently couple excitation light with substrates in glass-bottomed microwell plates, enabling rapid and precise detection of conformational changes induced by test entities.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If fluorescence-based detection is used for high throughput screening, then detection sensitivity is improved, but the system becomes sensitive to misalignment and instrumental drift requiring sophisticated optical systems
Solution Approach 1:
The patent replaces fluorescence-based optical detection with surface-selective second harmonic generation (SHG) detection. This substitution eliminates the need for sophisticated light sources, detectors, and optical alignment systems while maintaining high detection sensitivity. The SHG technique inherently provides surface selectivity and is less sensitive to instrumental drift and misalignment issues that plague fluorescence-based systems.
Solution Approach 2:
The patent changes the detection parameter from fluorescence emission to second harmonic generation signal. This parameter change transforms the detection mechanism to one that is surface-selective and less dependent on complex optical systems. The SHG signal arises from the non-linear optical response of the surface, providing inherent surface selectivity without requiring the sophisticated optical instrumentation needed for fluorescence detection.
2Measurement precision
If fluorescence-based detection is used for high throughput screening, then detection sensitivity is improved, but photo-bleaching causes signal degradation over time
Solution Approach 1:
The patent substitutes fluorescence detection with surface-selective second harmonic generation detection. This replacement eliminates photo-bleaching issues because SHG is a non-absorptive process that does not degrade the sample over time. The technique allows for repeated measurements on the same samples without signal degradation, enabling long-duration high throughput screening experiments.
3Reliability
If conventional small molecule or biologic drugs are used to block protein-protein interactions, then the target is inhibited, but the large contact area makes blocking extremely difficult
Solution Approach 1:
The patent applies local quality by using surface-selective detection to identify compounds that bind to specific local sites on the protein surface. Rather than attempting to block the entire large contact area of protein-protein interactions, the method identifies compounds that bind to particular regions or pockets on the protein surface, thereby inducing conformational changes that prevent the interaction. This localized approach makes drug discovery for large interface targets feasible.
Solution Approach 2:
The patent uses an intermediary approach by detecting conformational changes in the protein rather than directly measuring the protein-protein interaction. The surface-selective SHG detection serves as an intermediary readout that reports on protein conformational state. This allows identification of compounds that modulate protein conformation without requiring direct observation or blocking of the large interaction interface.
4Measurement precision
If X-ray crystallography or NMR methods are used to obtain structural information, then structural data is obtained, but throughput is low and sensitivity is limited
Solution Approach 1:
The patent replaces X-ray crystallography and NMR methods with surface-selective second harmonic generation detection. This substitution dramatically increases throughput while maintaining the ability to detect structural and conformational information. The SHG method can rapidly screen large numbers of samples in a high throughput format, unlike the low-throughput nature of crystallography and NMR.
Solution Approach 2:
The patent transitions from bulk structural analysis methods (X-ray, NMR) to surface-selective detection. This dimensional change focuses the measurement on the surface layer where the biological activity occurs, enabling rapid screening while capturing conformational information. The surface selectivity provides a new dimension of measurement that is both high throughput and sensitive to conformational changes.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This approach allows for rapid detection of conformational changes in biological entities at high throughput rates, overcoming the limitations of fluorescence-based methods and enabling the identification of allosteric modulators for 'undruggable' targets with reduced clinical side effects.
Implementation Method 1
The systems and methods disclosed herein provide mechanisms for coupling the high intensity excitation light required for SHG and other nonlinear optical techniques to a substrate, e.g. the glass substrate in a glass-bottomed microwell plate, by means of total internal reflection
Implementation Method 2
second harmonic generation (SHG) is a nonlinear optical process which may be configured as surface-selective detection technique that enables detection of conformational change in proteins and other biological targets
Data Source
Figure 1~1B
Figure 2
Figure 3~3D
AI summary
Methods, devices, and systems are disclosed for performing high throughput analysis of conformational change in biological molecules or other biological entities using surface-selective nonlinear optical detection techniques.