Recombinant Xaa-Pro dipeptidase (pepQ)
Description
Introduction to Recombinant Xaa-Pro Dipeptidase (pepQ)
Recombinant Xaa-Pro dipeptidase (pepQ), also known as prolidase or peptidase-Q, is a metalloenzyme belonging to the M24B family. It catalyzes the hydrolysis of dipeptides with a proline (Pro) or hydroxyproline (Hyp) residue at the C-terminus, playing roles in protein degradation, collagen recycling, and detoxification of organophosphorus compounds . First identified in Escherichia coli, pepQ is widely expressed in bacteria, archaea, and eukaryotes, with recombinant variants optimized for industrial and therapeutic applications .
2.1. Molecular Architecture
Recombinant pepQ exists as a homodimer, with each subunit containing:
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N-terminal domain: Involved in substrate binding.
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C-terminal domain: Contains the active site with a "pita-bread" fold, characteristic of the M24B family .
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Metal cofactors: Typically binds two Mn²⁺ ions, though Co²⁺ or Zn²⁺ variants are reported in some organisms .
| Property | Value | Source |
|---|---|---|
| Molecular weight | ~43–48 kDa (monomer) | |
| Optimal pH | 7.0–8.5 (varies by organism) | |
| Temperature stability | 25–37°C (optimal activity) |
2.2. Catalytic Mechanism
The mechanism involves:
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Metal coordination: Mn²⁺ ions activate a water molecule for nucleophilic attack on the scissile bond.
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Substrate binding: His255 coordinates the carboxylate group of the Pro residue, while Arg398 stabilizes the C-terminal oxygen .
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Tetrahedral intermediate: Formation via hydroxide attack, leading to peptide bond cleavage .
3.1. Substrate Preferences
pepQ exhibits broad specificity for Xaa-Pro dipeptides but shows higher activity toward hydrophobic residues (e.g., Met-Pro, Leu-Pro) .
| Substrate | k<sub>cat</sub> (s⁻¹) | K<sub>m</sub> (µM) | k<sub>cat</sub>/K<sub>m</sub> (M⁻¹s⁻¹) |
|---|---|---|---|
| Met-Pro | 109 | 0.13 | 8.4 × 10⁵ |
| Lys-Pro | 45.2 | 0.15 | 3.0 × 10⁵ |
| Gly-Pro | 22.1 | 0.12 | 1.8 × 10⁵ |
| Data from E. coli pepQ |
3.2. Metal Dependency
Activity is modulated by divalent cations:
4.1. Industrial and Biotechnological Uses
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Food processing: Reduces bitterness in cheese by hydrolyzing Pro-containing dipeptides .
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Organophosphate detoxification: Hydrolyzes nerve agents (e.g., sarin, soman) and pesticides .
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Collagen recycling: Critical for Pro reutilization in human metabolism .
