4D NMR Structure

Why to choose AI|ffinity’s 4D NMR-based method for protein structure determination?

Our new ML-based version, 4D-GRAPHS, enables automatic, all protein atom resonance assignment using only two 4D NMR spectra, reducing measurement time. For small proteins, a single 4D spectrum is sufficient. These resonance assignments enable 3D structure determination using standard tools.

Advantages:

• Suitable for flexible proteins: if they are soluble and stable at concentrations as low as 0.1-0.2 mM, with the typical range being 0.2 mM to 1 mM; non need for crystallization.
• Speed and Cost-Effectiveness: up to 6x faster and 3x cheaper than X-ray crystallography and up to 2x faster than standard 3D NMR methods.
• Handling Larger Proteins: thanks to the enhanced 4D peak resolution.
• Less Human Input: streamlined spectrum analysis and structure calculations reduce human intervention and related errors.
• Market-Leading Speed: only one 4D spectrum is needed for follow-up structures, such as mutants or protein-ligand complexes.
• Insight Into Protein Dynamics: and mechanisms of action.
• Mapping Interactions: with other molecules and is ideal for weak binders (hit compounds), offering a competitive edge.
• Natural Environment Representation: unlike X-ray crystallography and Cryo-EM that provide structural information about proteins in an artificially "frozen" state.
• No Installation Needed: the algorithm and all necessary software for structure calculations will be provided as a cloud services, including technical support.

Disadvantages:

• Protein Size Limit: at ~30 kDa, although this limit is expected to increase with new Ultra-high field NMR spectrometers (1.2 GHz).
• Dependence on Third-Party Software: for raw spectra processing, which can make the pipeline more complex, though this issue is common to all NMR software for resonance assignment.
• Requirement for High Protein Concentrations: typically 0.5 mM to 2 mM for globular proteins, with larger ones often requiring the upper end for adequate signal-to-noise ratios, risking potential aggregation or precipitation.

What are the comparative strengths and weaknesses of AI|ffinity's 4D NMR-based technique for protein structure determination versus other available experimental and computational methods?

In addition to 4D NMR structure determination, we also perform a comprehensive array of standard NMR experiments within our drug discovery projects.

Ligand-observe methods (1D NMR): focus on the ligand's spectral properties and have no target size limitations.

• binding validation
• ligand epitope mapping
• ligand epitope mapping
• binding constand (Kd) determination
• competition assays

Target-observe methods (2D NMR): focus on the protein's spectral properties and do have size limitations (~30 kDa).

• mapping of interactions with the protein
• protein conformational changes induced by ligand binding

Leveraging our 4D-NMR technology and computer-aided drug design (CADD), we're adept at designing potent inhibitors for biological targets. Trust AI|ffinity to accelerate your drug discovery process.