AnhE, a Metallochaperone Involved in the Maturation of a Cobalt-dependent Nitrile Hydratase
Bibliographic record
Abstract
Acetonitrile hydratase (ANHase) of Rhodococcus jostii RHA1 is a cobalt-containing enzyme with no significant sequence identity with characterized nitrile hydratases. The ANHase structural genes anhA and anhB are separated by anhE, predicted to encode an 11.1-kDa polypeptide. An anhE deletion mutant did not grow on acetonitrile but grew on acetamide, the ANHase reaction product. Growth on acetonitrile was restored by providing anhE in trans. AnhA could be used to assemble ANHase in vitro, provided the growth medium was supplemented with 50 μM CoCl2. Ten- to 100-fold less CoCl2 sufficed when anhE was co-expressed with anhA. Moreover, AnhA contained more cobalt when produced in cells containing AnhE. Chromatographic analyses revealed that AnhE existed as a monomer-dimer equilibrium (100 mm phosphate, pH 7.0, 25 °C). Divalent metal ions including Co2+, Cu2+, Zn2+, and Ni2+ stabilized the dimer. Isothermal titration calorimetry studies demonstrated that AnhE binds two half-equivalents of Co2+ with Kd of 0.12 ± 0.06 nm and 110 ± 35 nm, respectively. By contrast, AnhE bound only one half-equivalent of Zn2+ (Kd = 11 ± 2 nm) and Ni2+ (Kd = 49 ± 17 nm) and did not detectably bind Cu2+. Substitution of the sole histidine residue did not affect Co2+ binding. Holo-AnhE had a weak absorption band at 490 nm (ϵ = 9.7 ± 0.1 m−1 cm−1), consistent with hexacoordinate cobalt. The data support a model in which AnhE acts as a dimeric metallochaperone to deliver cobalt to ANHase. This study provides insight into the maturation of NHases and metallochaperone function. Acetonitrile hydratase (ANHase) of Rhodococcus jostii RHA1 is a cobalt-containing enzyme with no significant sequence identity with characterized nitrile hydratases. The ANHase structural genes anhA and anhB are separated by anhE, predicted to encode an 11.1-kDa polypeptide. An anhE deletion mutant did not grow on acetonitrile but grew on acetamide, the ANHase reaction product. Growth on acetonitrile was restored by providing anhE in trans. AnhA could be used to assemble ANHase in vitro, provided the growth medium was supplemented with 50 μM CoCl2. Ten- to 100-fold less CoCl2 sufficed when anhE was co-expressed with anhA. Moreover, AnhA contained more cobalt when produced in cells containing AnhE. Chromatographic analyses revealed that AnhE existed as a monomer-dimer equilibrium (100 mm phosphate, pH 7.0, 25 °C). Divalent metal ions including Co2+, Cu2+, Zn2+, and Ni2+ stabilized the dimer. Isothermal titration calorimetry studies demonstrated that AnhE binds two half-equivalents of Co2+ with Kd of 0.12 ± 0.06 nm and 110 ± 35 nm, respectively. By contrast, AnhE bound only one half-equivalent of Zn2+ (Kd = 11 ± 2 nm) and Ni2+ (Kd = 49 ± 17 nm) and did not detectably bind Cu2+. Substitution of the sole histidine residue did not affect Co2+ binding. Holo-AnhE had a weak absorption band at 490 nm (ϵ = 9.7 ± 0.1 m−1 cm−1), consistent with hexacoordinate cobalt. The data support a model in which AnhE acts as a dimeric metallochaperone to deliver cobalt to ANHase. This study provides insight into the maturation of NHases and metallochaperone function.
Fetched live from OpenAlex and de-inverted. Abstracts are not stored in this database: the inverted indexes are 8.6 GB of the frame’s 9.3 GB of text, and the host has 13 GB free.
How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.001 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 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 teacher head, 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".