9NLT

Crystal Structure of Flpp3-minibinder687 Complex

Summary for 9NLT

• Entry DOI: 10.2210/pdb9nlt/pdb
• Descriptor: DUF3568 family protein, minibinder687, IODIDE ION, ... (4 entities in total)
• Functional Keywords: de novo design, deep learning, plpp3, minibinder, francisella-like lipoprotein, apoptosis
• Biological source: Francisella tularensis; More
• Total number of polymer chains: 2
• Total formula weight: 18998.50

Authors

Bera, A.K.,Gokce, G.,Kang, A.,Bhardwaj, G. (deposition date: 2025-03-03, release date: 2025-10-22, Last modification date: 2025-11-05)

Primary citation

Gokce-Alpkilic, G.,Huang, B.,Liu, A.,Kreuk, L.S.M.,Wang, Y.,Adebomi, V.,Bueso, Y.F.,Bera, A.K.,Kang, A.,Gerben, S.R.,Rettie, S.,Vafeados, D.K.,Roullier, N.,Goreshnik, I.,Li, X.,Baker, D.,Woodward, J.J.,Mougous, J.D.,Bhardwaj, G.
De Novo Design of High-Affinity Miniprotein Binders Targeting Francisella Tularensis Virulence Factor.
Angew.Chem.Int.Ed.Engl., :e202516058-e202516058, 2025

PubMed Abstract: Francisella tularensis poses considerable public health risk due to its high infectivity and potential for bioterrorism. Francisella-like lipoprotein (Flpp3), a key virulence factor unique to Francisella, plays critical roles in infection and immune evasion, making it a promising target for therapeutic development. However, the lack of well-defined binding pockets and structural information on native interactions has hindered structure-guided ligand discovery against Flpp3. Here, we used a combination of physics-based and deep-learning methods to design high-affinity miniprotein binders targeting two distinct sites on Flpp3. We identified four binders for site I with binding affinities ranging between 24-110 nM. For the second site, an initial binder showed a dissociation constant (K) of 81 nM, and subsequent site saturation mutagenesis yielded variants with sub-nanomolar affinities. Circular dichroism confirmed the topology of designed miniproteins. The X-ray crystal structure of Flpp3 in complex with a site I binder is nearly identical to the design model (Cα root-mean-square deviation (RMSD): 0.9 Å). These designed miniproteins provide research tools to explore the roles of Flpp3 in tularemia and should enable the development of new therapeutic candidates.

PubMed: 41117072
DOI: 10.1002/anie.202516058

Experimental method

X-RAY DIFFRACTION (2.37 Å)