The putative type I secretion sequence of a 1.5-MDa icebinding adhesin folds as a Ca2+-dependent β-rich structure
Bibliographic record
Abstract
Marinomonas primoryensis is a Gram-negative and strictly aerobic bacterium, an isolate of which was found in a brackish (half-strength seawater) lake in Antarctica. M. primoryensis produces a Ca2+-dependent 1.5-MDa ice-binding adhesin (MpAFP) that likely binds the bacterium to the underside of surface ice, where it may have better access to oxygen and other nutrients. The MpAFP is divided into five distinct regions that include the highly repetitive Region II (RII) and the moderately repetitive ice-binding Region IV (RIV). Region V (RV) is the C-terminal domain of MpAFP, and may serve as its non-cleavable signal sequence for the Type I Secretion System (TISS). The Protein Homology/analogY Recognition Engine (Phyre2) Server modeled RV as a Ca2+-bound β-rich structure. A previous study used a RV construct that started directly following the C-terminus of RIV, but the expressed protein aggregated and was unsuitable for further characterization. Here, we report the expression and purification of a fusion construct spanning RIV and RV. We defined the start of the RV domain by digesting the purified RIV-V fusion protein with trypsin in the presence of Ca2+. A soluble RV was then purified from the digestion mixture using anion-exchange chromatography, and its N-terminal residues were determined using Edman degradation sequencing. The folding of the newly designed RV construct was assessed by size-exclusion chromatography and circular dichroism (CD) spectroscopy. This work will provide insight into the structural relationship between RIV and RV, as well as how extremely large proteins are secreted via the TISS in Gram-negative bacteria.
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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 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.001 |
| 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 | 0.001 |
| Insufficient payload (model declined to judge) | 0.001 | 0.001 |
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".