Human Plasma Membrane Anion Exchanger, AE1: Over‐expression in <i>Saccharomyces cerevisiae</i>
Bibliographic record
Abstract
Anion exchanger 1 (AE1) facilitates the exchange of chloride for bicarbonate across the plasma membrane of renal and erythrocyte membranes. The 55 kDa membranedomain of AE1 alone is responsible for its transport function, however a high resolution structure of this domain has not been solved. The limiting factor to obtaining acrystal structure is the availability of homogenous, purified AE1. Here we developed a novel expression and efficient purification system for AE1, using S. cerevisiae to allow for large‐scale expression, while maintaining proper folding of AE1. The expression construct consists of the AE1 membrane domain (AE1MD), residues 388–911 of human AE1, followed by the 9 C‐terminal amino acids of rhodopsin. Expression was under the control of the constitutive promoter and transcriptionalterminator of the yeast H + ‐ATPase gene. AE1MD was expressed in S. cerevisiae, strain BJ5457, to 0.3 mg AE1MD per litre of culture. We characterized the cellularlocalization of AE1MD by confocal immunofluorescence microscopy and sucrose gradient ultracentrifugation. AE1MD structure was assessed by binding to animmobilized anion exchange inhibitor (SITS‐Affi‐gel). Once solubilized by the detergent n‐dodecyl‐β‐D‐maltoside, AE1MD was purified with a 1D4 antibody affinitycolumn, which recognizes the rhodopsin tag on AE1MD. This research is supported by the Canadian Institutes of Health Research.
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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 | 0.000 | 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".