The design of enzymes with complex active sites that mediate multistep reactions remains an outstanding challenge. With serine hydrolases as a model system, we combined the generative capabilities of RFdiffusion with ...
The design of enzymes with complex active sites that mediate multistep reactions remains an outstanding challenge. With serine hydrolases as a model system, we combined the generative capabilities of RFdiffusion with an ensemble generation method for assessing active site preorganization at each step in the reaction to design enzymes starting from minimal active site descriptions. Experimental characterization revealed catalytic efficiencies (kcat/Km) up to 2.2 × 105 M-1 s-1 and crystal structures that closely match the design models (Cα root mean square deviations <1 angstrom). Selection for structural compatibility across the reaction coordinate enabled identification of new catalysts remove with five different folds distinct from those of natural serine hydrolases. Our de novo approach provides insight into the geometric basis of catalysis and a roadmap for designing enzymes that catalyze multistep transformations.
Correction to: Nature Biotechnologyhttps://***/10.1038/s41587-024-02395-w, published online 25 September 2024. In the version of the article initially published, in the key to Fig. 2f the colors were swapped and have ...
The existence of broadly cross-reactive antibodies that can neutralize diverse HIV-1 isolates (bnAbs) has been appreciated for more than a decade. Many high-resolution structures of bnAbs, typically with one or two we...
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The existence of broadly cross-reactive antibodies that can neutralize diverse HIV-1 isolates (bnAbs) has been appreciated for more than a decade. Many high-resolution structures of bnAbs, typically with one or two well-characterized HIV-1 Env glycoprotein trimers, have been reported. However, an understanding of how such antibodies grapple with variability in their antigenic targets across diverse viral isolates has remained elusive. To achieve such an understanding requires first characterizing the extent of structural and antigenic variation embodied in Env, and then identifying how a bnAb overcomes that variation at a structural level. Here, using hydrogen/deuterium-exchange mass spectrometry (HDX-MS) and quantitative measurements of antibody binding kinetics, we show that variation in structural ordering in the V1/V2 apex of Env across a globally representative panel of HIV-1 isolates has a marked effect on antibody association rates and affinities. We also report cryo-EM reconstructions of the apex-targeting PGT145 bnAb bound to two divergent Env that exhibit different degrees of structural dynamics throughout the trimer structures. Parallel HDX-MS experiments demonstrate that PGT145 bnAb has an exquisitely focused footprint at the trimer apex where binding did not yield allosteric changes throughout the rest of the structure. These results demonstrate that structural dynamics are a cryptic determinant of antigenicity, and mature antibodies that have achieved breadth and potency in some cases are able to achieve their broad cross-reactivity by "threading the needle" and binding in a highly focused fashion, thus evading and overcoming the variable properties found in Env from divergent isolates.
We use computational design coupled with experimental characterization to systematically investigate the design principles for macrocycle membrane permeability and oral *** designed 1846-12 residue macrocycles with a ...
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We use computational design coupled with experimental characterization to systematically investigate the design principles for macrocycle membrane permeability and oral *** designed 1846-12 residue macrocycles with a wide range of predicted structures containing noncanonical backbone modifications and experimentally determined structures of 35;29 are very close to the computational *** such control,we show that membrane permeability can be systematically achieved by ensuring all amide(NH)groups are engaged in internal hydrogen bonding interactions.84 designs over the 6-12 residue size range cross membranes with an apparent permeability greater than 1×10^(-6)cm/s.
作者:
Long GuiEric JurgensJamie L EbnerMatteo PorottoAnne MosconaKelly K LeeDepartment of Medicinal Chemistry
University of Washington Seattle Washington USA Biological Physics Structure and Design Graduate Program University of Washington Seattle Washington USA. Department of Pediatrics
Weill Medical College of Cornell University New York New York USA. Department of Medicinal Chemistry
University of Washington Seattle Washington USA. Department of Pediatrics
Weill Medical College of Cornell University New York New York USA kklee@uw.edu Anm2047@med.cornell.edu Map2028@med.cornell.edu. Department of Pediatrics
Weill Medical College of Cornell University New York New York USA Department of Microbiology and Immunology Weill Medical College of Cornell University New York New York USA kklee@uw.edu Anm2047@med.cornell.edu Map2028@med.cornell.edu. Department of Medicinal Chemistry
University of Washington Seattle Washington USA Biological Physics Structure and Design Graduate Program University of Washington Seattle Washington USA Department of Microbiology University of Washington Seattle Washington USA kklee@uw.edu Anm2047@med.cornell.edu Map2028@med.cornell.edu.
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