biostudies-literatureSudhamalla BUniversity of PittsburghNational Institute of General Medical SciencesNIGMS NIH HHS10263-10269https://www.ebi.ac.uk/biostudies/studies/S-EPMC6400064biostudies-literatureUnknown140(32)Ten-eleven translocation (TET) enzymes employ O₂, earth-abundant iron, and 2-ketoglutarate (2KG) to perform iterative C-H oxidation of 5-methylcytosine in DNA to control expression of the mammalian genome. Given that more than 60 such C-H oxygenases are present in humans, determining context-dependent functions of each of these enzymes is a pivotal challenge. In an effort to tackle the problem, we developed analogue-sensitive TET enzymes to perturb the activity of a specific member. We rationally engineered the TET2-2KG interface to develop TET2 variants with an expanded active site that can be specifically inhibited by the N-oxalylglycine (NOG) derivatives carrying a complementary steric "bump". Herein, we describe the identification and engineering of a bulky gatekeeper residuJournal of the American Chemical SocietyComplementary Steric Engineering at the Protein-Ligand Interface for Analogue-Sensitive TET Oxygenases.PMC6400064R01GM123234R01 GM123234Kavoosi SSnyder VWang SIslam KSudhamalla BArora SfalseComplementary Steric Engineering at the Protein-Ligand Interface for Analogue-Sensitive TET Oxygenases.Ten-eleven translocation (TET) enzymes employ O₂, earth-abundant iron, and 2-ketoglutarate (2KG) to perform iterative C-H oxidation of 5-methylcytosine in DNA to control expression of the mammalian genome. Given that more than 60 such C-H oxygenases are present in humans, determining context-dependent functions of each of these enzymes is a pivotal challenge. In an effort to tackle the problem, we developed analogue-sensitive TET enzymes to perturb the activity of a specific member. We rationally engineered the TET2-2KG interface to develop TET2 variants with an expanded active site that can be specifically inhibited by the N-oxalylglycine (NOG) derivatives carrying a complementary steric "bump". Herein, we describe the identification and engineering of a bulky gatekeeper residu2018-01-01T00:00:00Z2018 Aug2026-05-05T06:28:00.586Z2025-05-29T20:01:53.177ZS-EPMC64000643002860010.1021/jacs.8b05283