Single-Domain Metallothioneins: Evidence of the Onset of Clustered Metal Binding Domains in Zn-rhMT 1a
Bibliographic record
Abstract
Mammalian metallothioneins bind up to seven Zn(2+) ions in two distinct domains: an N-terminal β-domain that binds three Zn(2+) ions and a C-terminal α-domain that binds four Zn(2+) ions. Domain specificity has been invoked in the metalation mechanism with cluster formation and bridging of the 20 Cys residues taking place prior to saturation with seven Zn(2+) ions. We report a novel experiment that examines Zn(2+) metalation by exploiting the expected decrease in K(F) at the onset of clustering using electrospray ionization mass spectrometry (ESI-MS). During the titration with Zn(2+), the ESI-MS data show that several metalated species coexist until the fully saturated proteins are formed. The relative Zn binding affinities of the seven total sites in the α- and β-fragments were determined through direct competition for added Zn(2+). The K(F) values for each Zn(2+) are expected to decrease as a function of the remaining available sites and the onset of clustering. Analysis shows that Zn(2+) binds to β-rhMT with a greater affinity than α-rhMT. The incremental distribution of Zn(2+) between the competing fragments and apo-βα-rhMT (essentially three and four sites competing with seven sites) identifies the exact point at which clustering begins in the full protein. Analysis of the speciation data shows that Zn(5)-MT forms before clustering begins. This means that all 20 Cys residues of apo-βα-rhMT are bound terminally to Zn(2+) as [Zn(Cys)(4)](2-) units before clustering begins; there is no domain preference in this first metalation stage. Preferential binding of Zn(2+) to β- and α-rhMT at the point where βα-rhMT must form clusters is caused by a significant decrease in the affinity of βα-rhMT for further Zn(2+). The single-domain Zn(5)-rhMT, in which there are no exposed cysteine sulfurs, is a key component of the metalation pathway because the lower affinities of the two clustered Zn(2+) ions allow donation to apoenzymes.
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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 | 0.001 |
| 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 | 0.001 | 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".