Galectin-3 Binding Protein Contaminates the Purification of Recombinant Factor IX.
Bibliographic record
Abstract
Abstract The hemophilas are the most common severe hereditary bleeding disorders. The standard treatment of these disorders is to infuse plasma concentrates of the missing coagulation factor. However, the high cost of replacement therapy and the serious morbidity from the transmission of infectious agents have mandated a need for alternative therapeutic strategies. One of these strategies is gene therapy and we embarked on a novel gene therapeutic strategy involving the use of factor IX-transduced bone marrow stroma to treat hemopihlia B, a deficiency of the gamma-carboxylated protein, factor IX. The rate limiting step for the production of factor IX is gamma-carboxylation, a post translational modification carried out only in mammalian cells. In order to test the carboxylation efficiency of our gene therapeutic platform, we first transfected Human Embryonic Kidney (HEK) 293 cells with a retroviral construct containing factor IX. Factor IX was purified by immunoaffinity chromatography and then subject to hydroxyapatite chromatography to isolate biologically active properly carboxylated factor IX from inactive uncarboxylated factor IX. Unexpectedly, during hydroxyapatite chromatography, we discovered that purified factor IX was contaminated by a heretofore unknown protein. Further analysis by mass spectrometry sequencing revealed this protein to be galectin-3 binding protein (G3BP).G3BP contaminates media of HEK293 cells regardless of whether factor IX is transfected into these cells. Furthermore, the contamination of purified media is not specific for factor IX as G3BP is isolated regardless of the antibody used for purification. In summary, the above results provide important information regarding the production of recombinant factor IX and uncover a potentially new contaminant in recombinant products produced in mammalian cells.
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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) | 0.001 | 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 | 0.001 |
| Insufficient payload (model declined to judge) | 0.002 | 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".