Evidence for the Amphipathic Nature and Tilted Topology of Helices 4 and 5 in the Closed State of the Colicin E1 Channel
Bibliographic record
Abstract
The membrane-bound closed channel structure of helices 4 and 5 (Lys-406-Asp-446) of the colicin E1 channel domain was investigated by using a site-directed fluorescence labeling technique. A bimane probe was covalently attached to a cysteine residue in a series of single-cysteine mutant proteins to scan each helix in a residue-by-residue fashion. A variety of fluorescence properties of the bimane fluorophore were measured for both the soluble and membrane-bound states of the channel peptide, including the fluorescence emission maximum, fluorescence anisotropy, and membrane bilayer penetration depth. The fluorescence properties were collated for construction of a membrane-bound topology model of helices 4 and 5 of the channel domain. Finally, the data reveal that both helices 4 and 5 are two separate amphipathic alpha-helices that are situated parallel to the membrane surface. Dual fluorescence quencher analysis shows that helix 4 adopts a tilted topology in which its C-terminus is more buried than its N-terminus within the bilayer. In contrast, helix 5 is relatively solvent-exposed and located parallel to the interfacial region of the membrane surface. However, the loop region of both helices 4 and 5 was shown to be relatively buried within the bilayer. In addition, a least-squares fit of the data to a harmonic wave function indicated that both periodicity and frequency are typical for an amphipathic alpha-helix (3.75 +/- 0.1 residues per turn for helix 4 and 3.70 +/- 0.1 residues per turn for helix 5). Squared residual plots of the harmonic wave function also reveal that the N-terminus of helix 4 is elongated by two residues from Lys-406 to Phe-404, while the C-terminus of helix 5 is elongated by three residues from Tyr-434 to Ile-437 upon membrane association.
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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 | 0.001 |
| 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 | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.001 | 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".