26AP

Complex between N-lobe Arc mutant F267/F5Phe and nanobody H11

Summary for 26AP

• Entry DOI: 10.2210/pdb26ap/pdb
• Descriptor: Nanobody H11, Activity-regulated cytoskeleton-associated protein, DI(HYDROXYETHYL)ETHER, ... (4 entities in total)
• Functional Keywords: antibody-antigen complex, non-canonical amino acids, pentafluorophenylalanine, immune system
• Biological source: Vicugna pacos; More
• Total number of polymer chains: 2
• Total formula weight: 24179.64

Authors

Macri, L.M.,Habel, E.,Huber, T. (deposition date: 2026-04-24, release date: 2026-08-05, Last modification date: 2026-08-12)

Primary citation

Paul, N.,Welegedara, A.P.,Frkic, R.L.,Macri, L.,Thompson, T.R.C.,Baber, J.L.,Habel, E.,Abdelkader, E.H.,Qianzhu, H.,Chilton, N.F.,Jackson, C.J.,Bax, A.,Huber, T.,Otting, G.
Genetically Encoded Pentafluorophenylalanine Enables Quantitative Probing of Local Protein Malleability by 19 F NMR.
J.Am.Chem.Soc., 2026

PubMed Abstract: Aromatic ring flips in proteins provide a direct probe of local structural fluctuations, yet their rates are typically too fast for quantitative measurement by NMR spectroscopy. Here we show that site-specific incorporation of 2,3,4,5,6-pentafluoro-l-phenylalanine (F5Phe) reshapes the torsional energy landscape of aromatic side chains, slowing ring flips by over 2 orders of magnitude and shifting them into the slow-exchange regime accessible by 19F NMR. F5Phe can be genetically encoded with high fidelity and minimal structural perturbation, as confirmed by high-resolution X-ray crystallography across multiple proteins. The resulting 19F NMR spectra enable direct, quantitative measurements of ring-flip kinetics without the need for isotope labeling or complex multidimensional experiments. Application to a diverse set of proteins demonstrates that ring-flip rates vary widely even within the same hydrophobic cluster, revealing highly localized conformational fluctuations rather than global unfolding events. Pressure-dependent measurements yield small activation volumes, indicating that the structural rearrangements enabling ring flips are spatially confined. Ligand binding and protein-protein interactions modulate ring-flip rates in a site-specific manner, providing a sensitive readout of allosteric effects on local protein malleability. These results establish fluorinated aromatic amino acids as a general chemical strategy to engineer dynamic observables in proteins, transforming aromatic ring flips into a broadly applicable probe of local conformational dynamics and allostery.

PubMed: 42503665
DOI: 10.1021/jacs.6c10121

Experimental method

X-RAY DIFFRACTION (1.5 Å)