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@article{pmid42935473,

title = {Site-Selective Bioconjugations of Distinct Methionine Residues in a Protein via Reactivity-Based Reagent Control},

author = {Elizabeth J Sisko and Kaan Kumru and James A Wells and Jason K Sello},

doi = {10.1021/acschembio.6c00636},

issn = {1554-8937},

year  = {2026},

date = {2026-10-27},

urldate = {2026-10-01},

journal = {ACS Chem Biol},

abstract = {The reaction of thioethers in biological macromolecules with oxaziridines enables their functionalization with moieties having utility in both basic and applied science. Exploitation of that reaction has advantages over common bioconjugations due to the relatively low abundance and weak reactivity of thioethers. Herein, we report a novel and improved route to 1-ureal piperidine,3-aryl oxaziridines that enables the preparation of bioconjugation reagents that are either electronically activated or deactivated. We leveraged the differential reactivities of the oxaziridine reagents and engineered methionine residues in distinct environments within a therapeutically relevant antibody fragment (Fab) to effect site-selective bioconjugations. We capitalized on this novel means of kinetic resolution to effect dual functionalization of a HER2-targeted Fab with distinct fluorophores for imaging receptor internalization. This work broadens the scope of what is possible in bioconjugations of thioethers and demonstrates that site-selective bioconjugations of the same residue at more than one site in a protein can be realized via reactivity-based reagent control.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid42853246,

title = {Structural basis of serine protease inhibition by antibodies from biased Fab phage-display libraries},

author = {Kyle J Anderson and Melody S Lee and Natalia Sevillano and Gang Chen and Michael J Hornsby and Sachdev S Sidhu and Charles S Craik},

doi = {10.1515/hsz-2026-0121},

issn = {1437-4315},

year  = {2026},

date = {2026-10-01},

urldate = {2026-10-01},

journal = {Biol Chem},

abstract = {Biased Fab phage-display libraries were designed to determine whether inhibitory CDR H3 motifs from potent anti-matriptase antibodies could be transferred to target homologous serine proteases. Using reverse-binding and substrate-like H3 motifs from parental clones A11 and E2 as templates, six synthetic libraries with 10 diversity were constructed. Selection against matriptase identified 16 inhibitors with sub-100 nM potency, representing 100,000-fold improvement over circularized H3 loops alone. Selection against TMPRSS2, a serine protease implicated in viral entry and prostate cancer with 43 % sequence identity to matriptase, yielded binders with micromolar inhibitory potency. Selection against urokinase plasminogen activator (uPA, 35 % identity) identified binders that adopted a substrate-like CDR H3 binding mode in our structural models. Across all reference structures, including the separately identified uPA inhibitor AB2 (PDB: 9PYF, deposited with this work), benchmarking of five co-folding methods and rigid-body docking showed that co-folding consistently achieved acceptable to high quality DockQ scores, outperforming traditional docking and capturing the recognition of key active site determinants. Ensemble predictions of mutational binding energy changes (ΔΔG) using these models identified key paratope-epitope interactions, with predictions validated through mutagenesis. This work establishes a framework integrating biased antibody libraries with computational structure prediction and analysis for targeting conserved protease epitopes.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}

@article{pmid42905542,

title = {Comparative analysis of photocatalytic, enzymatic, and chemical approaches to identify canonical and noncanonical surfaceomes},

author = {William W Ho and Yifei Chen and Tomasz Slezak and Oleksandr Zginnyk and Trenton M Peters-Clarke and Kaitlin Schaefer and Anthony A Kossiakoff and Kevin K Leung and James A Wells},

doi = {10.1073/pnas.2623700123},

issn = {1091-6490},

year  = {2026},

date = {2026-10-01},

urldate = {2026-10-01},

journal = {Proc Natl Acad Sci U S A},

volume = {123},

number = {43},

pages = {e2623700123},

abstract = {The cell surface is a critical nexus in cellular communication and a rich landscape yielding the majority of current therapeutic targets, yet only a small fraction of cell surface proteins have been targeted therapeutically. The cell surface proteome (surfaceome) comprises only ~1% of the human proteome by mass, motivating enrichment strategies to detect surfaceome alterations in disease for drug target discovery. Developments in photocatalytic technology have revolutionized the study of protein neighborhoods around specific targets of interest, but photocatalysis has been underutilized for global surfaceome analysis. Here, we describe five distinct photocatalytic surfaceomics strategies, all without the need for cellular engineering, that apply the photocatalyst Eosin Y attached to different cell surface structures: direct glycan ligation, glycan attachment via two different lectins, membrane insertion, and free in extracellular space. We systematically compare these photocatalytic surfaceomics methods to each other as well as to three established chemical and enzymatic surfaceomics methods. We demonstrate that the different photocatalytic surfaceomics techniques achieve broadly similar canonical cell surface protein enrichment but show some important differences reflective of cell surface attachment mechanism. These mechanistically orthogonal surfaceomics techniques consistently identified numerous nonglycosylated proteins with high concordance, many of which represent an underappreciated class of noncanonical cell surface proteins lacking traditional cell surface protein features that may represent potential drug targets. This study substantially expands options for surfaceomics methods using photocatalytic proximity labeling.},

keywords = {},

pubstate = {published},

tppubtype = {article}

}