Structural and mechanistic views of enzymatic, heme-dependent nitrogen-nitrogen bond formation
Bibliographic record
Abstract
Summary Molecules with nitrogen-nitrogen (N-N) bonds constitute a large group of clinically important drugs, and various synthetic approaches have been developed to construct functional groups like hydrazines, diazos, pyrazoles, and N-nitrosos. While hundreds of N-N-containing specialized natural metabolites have also been discovered, little is known about the underlying enzymatic mechanisms that have evolved for N-N bond formation. In order to directly form a single N(sp 3 )-N(sp 3 ) bond, enzymes must reverse the typical nucleophilicity of one nitrogen. Here, we report structural and mechanistic interrogations of the piperazate synthase PipS, a heme-dependent enzyme that catalyzes an N-N bond forming cyclization of N 5 -OH-L-ornithine to give the non-proteinogenic amino acid L-piperazic acid. We show that PipS can process a variety of N -substituted hydroxylamines, to give either an imine or an N-N bond, in a substrate-specific manner. Using a combination of structural and biochemical experiments, computational studies, and spectroscopic characterization, we propose that heme-dependent dehydration and N-N bond formation in PipS proceed through divergent pathways, which may stem from a shared nitrenoid intermediate that effectively reverses the nucleophilicity of the hydroxylamine nitrogen. Our work expands the current knowledge of enzymatic N-N bond formation, and delineates the catalytic versatility of a heme cofactor, paving the way for future development of genetically encoded biocatalysts for N-N bond formation.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.001 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.001 |
| Scholarly communication | 0.001 | 0.001 |
| Open science | 0.001 | 0.000 |
| Research integrity | 0.001 | 0.001 |
| Insufficient payload (model declined to judge) | 0.002 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".