All Publications


  • Formylglycine-generating enzyme binds substrate directly at a mononuclear Cu(I) center to initiate O2 activation. Proceedings of the National Academy of Sciences of the United States of America Appel, M. J., Meier, K. K., Lafrance-Vanasse, J., Lim, H., Tsai, C., Hedman, B., Hodgson, K. O., Tainer, J. A., Solomon, E. I., Bertozzi, C. R. 2019

    Abstract

    The formylglycine-generating enzyme (FGE) is required for the posttranslational activation of type I sulfatases by oxidation of an active-site cysteine to Calpha-formylglycine. FGE has emerged as an enabling biotechnology tool due to the robust utility of the aldehyde product as a bioconjugation handle in recombinant proteins. Here, we show that Cu(I)-FGE is functional in O2 activation and reveal a high-resolution X-ray crystal structure of FGE in complex with its catalytic copper cofactor. We establish that the copper atom is coordinated by two active-site cysteine residues in a nearly linear geometry, supporting and extending prior biochemical and structural data. The active cuprous FGE complex was interrogated directly by X-ray absorption spectroscopy. These data unambiguously establish the configuration of the resting enzyme metal center and, importantly, reveal the formation of a three-coordinate tris(thiolate) trigonal planar complex upon substrate binding as furthermore supported by density functional theory (DFT) calculations. Critically, inner-sphere substrate coordination turns on O2 activation at the copper center. These collective results provide a detailed mechanistic framework for understanding why nature chose this structurally unique monocopper active site to catalyze oxidase chemistry for sulfatase activation.

    View details for DOI 10.1073/pnas.1818274116

    View details for PubMedID 30824597

  • Piperidine-based glycodendrons as protein N-glycan prosthetics BIOORGANIC & MEDICINAL CHEMISTRY Hudak, J. E., Belardi, B., Appel, M. J., Solania, A., Robinson, P. V., Bertozzi, C. R. 2016; 24 (20): 4791-4800

    Abstract

    The generation of homogeneously glycosylated proteins is essential for defining glycoform-specific activity and improving protein-based therapeutics. We present a novel glycodendron prosthetic which can be site-selectively appended to recombinant proteins to create 'N-glycosylated' glycoprotein mimics. Using computational modeling, we designed the dendrimer scaffold and protein attachment point to resemble the native N-glycan architecture. Three piperidine-melamine glycodendrimers were synthesized via a chemoenzymatic route and attached to human growth hormone and the Fc region of human IgG. These products represent a new class of engineered biosimilars bearing novel glycodendrimer structures.

    View details for DOI 10.1016/j.bmc.2016.05.050

    View details for Web of Science ID 000385905800005

    View details for PubMedID 27283789

    View details for PubMedCentralID PMC5052108