Kinetic Studies of HPr, HPr(H15D), HPr(H15E), and HPr(His∼P) Phosphorylation by the <i>Streptococcus salivarius</i> HPr(Ser) Kinase/Phosphorylase
Bibliographic record
Abstract
HPr is a central protein of the phosphoenolpyruvate:sugar phosphotransferase transport system (PTS). In streptococci, HPr can be phosphorylated at His(15) at the expense of PEP by enzyme I (EI) of the PTS, producing HPr(His approximately P). HPr can also be phosphorylated at Ser(46) by the ATP-dependent HPr(Ser) kinase/phosphorylase (HprK/P), producing HPr(Ser-P). Lastly, HPr can be phosphorylated on both residues, producing HPr(Ser-P)(His approximately P) (HPr-P2). We report here a study on the phosphorylation of Streptococcus salivarius HPr, HPr(H15D), HPr(H15E), and HPr(His approximately P) by HprK/P to assess the involvement of HprK/P in the synthesis of HPr-P2 in streptococcal cells. We first developed a spectrophotometric method for measuring HprK/P kinase activity. Using this assay, we found that the K(m) of HprK/P for HPr at pH 7.4 and 37 degrees C was approximately 110 muM, with a specificity constant (k(cat)/K(m)) of 1.7 x 10(4) M(-1) s(-1). The specificity constants for HPr(H15D) and HPr(H15E) were approximately 13 times lower. Kinetic studies conducted under conditions where HPr(His approximately P) was stable (i.e., pH 8.6 and 15 degrees C) showed that HPr(His approximately P) was a poorer substrate for HprK/P than HPr(H15D), the k(cat)/K(m) for HPr(H15D) and HPr(His approximately P) being approximately 9 and 26 times lower than that for HPr, respectively. Our results suggested that (i) the inefficiency of the phosphorylation of HPr(His approximately P) by HprK/P results from the presence of a negative charge at position 15 as well as from other structural elements and (ii) the contribution of streptococcal HprK/P to the synthesis of HPr-P2 in vivo is marginal.
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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 | 0.001 | 0.001 |
| Meta-epidemiology (narrow) | 0.001 | 0.000 |
| Meta-epidemiology (broad) | 0.001 | 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".