Anru Tian

All Publications

• Inverse FoldDir: Structure-conditioned Protein Sequence Design by Dirichlet Flow Matching. bioRxiv : the preprint server for biology Tartici, A., Stojkovic, M., Tian, A., Jewett, M. C., Altman, R. B., Wittmann, B. J. 2026

  Abstract

  Protein engineering has important implications in the bioeconomy, enabling applications in materials, medicine, and energy. A key challenge is designing protein sequences that have a specific form and function. Protein inverse folding seeks to address this challenge by identifying amino acid sequences compatible with a desired protein backbone. This task is central to protein redesign and can provide a sequence-design capability for de novo backbones produced by structure-generation methods. Ideally, inverse folding can provide diverse sequence alternatives, fixed residues or motifs, soft biochemical preferences at selected positions, and candidates that remain experimentally useful. We developed Inverse FoldDir, a controllable inverse-folding method that performs iterative denoising on the amino acid probability simplex. Given a backbone structure, the model updates all positions jointly through a learned Dirichlet flow, supporting full sequence generation, fixed-residue inpainting, and user-defined soft residue priors. On the held-out CATH 4.2 test set, Inverse FoldDir achieved a mean TM-score of 84.5 (on a 0-100 scale) and a mean Cα RMSD of 1.76 Å, compared with 83.3 and 1.86 Å, respectively, for ESM-IF1, the strongest evaluated baseline on both metrics. Denoising trajectory analyses showed that positions commit at different rates and that some residues change identity late in generation, illustrating whole-sequence refinement rather than one-shot prediction or irreversible sequential decoding. We experimentally tested Inverse FoldDir in an anti-GFP nanobody redesign task, where two of 35 redesigned sequences retained reproducible sfGFP-binding signal across independent assay runs with approximately 43% sequence divergence from the native nanobody. Inverse FoldDir is a structure-conditioned protein redesign method that combines structural recovery, user control, experimental validation, and a natural route toward future property-guided sampling.

  View details for DOI 10.64898/2026.09.06.749733

  View details for PubMedID 42818576

  View details for PubMedCentralID PMC13622493

• Topological reprogramming transforms an integral membrane oligosaccharyltransferase into a water-soluble glycosylation catalyst. bioRxiv : the preprint server for biology Kwon, Y. H., Mihaljević, L., Kim, K., Kim, D. E., Donahue, T. C., Bidstrup, E. J., Bandi, C. K., Sotomayor, B., Hulbert, S. W., Myers, K. A., Tian, A., Culpepper, M., Mizrachi, D., Jaroentomeechai, T., Clausen, H., Jewett, M. C., Baker, D., DeLisa, M. P. 2026

  Abstract

  Glycosyltransferases (GTs) catalyze the formation of new glycosidic bonds and thus are vital for synthesizing nature's vast repertoire of glycans and glycoconjugates and for engineering glycan-related medicines and materials. However, obtaining detailed structural and functional insights for the >750,000 known GTs is limited by difficulties associated with their efficient recombinant expression. Members of the GT-C fold, in particular, pose the most significant expression challenges due to the integration and folding requirements of their multiple membrane-spanning regions. Here, we address this challenge by engineering water-soluble variants of an archetypal GT-C fold enzyme, namely the oligosaccharyltransferase PglB from Campylobacter jejuni (CjPglB), which possesses 13 hydrophobic transmembrane helices. To render CjPglB water-soluble, we leveraged two advanced protein engineering methods: one that is universal called SIMPLEx (solubilization of IMPs with high levels of expression) and the other that is custom tailored called WRAPs (water-soluble RFdiffused amphipathic proteins). Each approach was able to transform CjPglB into a water-soluble enzyme that could be readily expressed in the cytoplasm of Escherichia coli cells at yields in the 3-6 mg/L range. Importantly, solubilization was achieved without the need for detergents and with retention of catalytic function. Collectively, our findings demonstrate that both SIMPLEx and WRAPs are promising platforms for advancing the molecular characterization of even the most structurally complex GTs, while also enabling broader GT-mediated glycosylation capabilities within synthetic glycobiology applications.

  View details for DOI 10.64898/2026.01.30.702934

  View details for PubMedID 41659604

  View details for PubMedCentralID PMC12879670