PURIFICATION AND CHARACTERIZATION OF THE 20S PROTEASOME FROM THEALGA <i>CHARA CORALLINA</i> (CHAROPHYCEAE)<sup>1</sup>
Bibliographic record
Abstract
We purified the 20S proteasome from the alga Chara corallina Willd with DEAE–ion‐exchange column chromatography and preparative nondenaturing PAGE. The analysis of the purified enzyme bynondenaturing PAGE gave a single band whose molecular mass was estimated to be about 600,000 Da by gel permeation chromatography and whose isoelectric point was at pH 5.5. Two‐dimensional gel electrophoresis gave at least 12 spots with molecular masses from 26,000 to 32,000 Da in a wide range of isoelectric points. The 20S proteasome hydrolyzed three types of artificial substrates used to differentiate chymotrypsin‐like, trypsin‐like, and peptidyl glutamyl peptidase activities. Both the chymotrypsin‐like and the peptidyl glutamyl peptidase activities were enhanced by SDS. In the presence of 0.03% SDS, the optimal pH for both activities was 8.5. Trypsin‐like activity of the 20S proteasome had a broad pH optimum in an alkaline region and was not activated but inhibited by SDS. Its chymotrypsin‐like activity was inhibited by N ‐ethylmaleimide, p ‐chloromercuribenzoic acid, and chymostatin. In contrast, its peptidyl glutamyl peptidase activity was not inhibited by chymostatin. Moreover, proteasome inhibitors MG 115 and MG 135 were effective against the chymotrypsin‐like activity and less so against the peptidyl glutamyl peptidase activity. These properties were very similar to those of the proteasomes of mammalian, yeast, and spinach cells. The large size of Chara cells will make in vivo manipulations and investigations of the proteasome proteolytic system possible.
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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".