Recombinant Chlamydomonas reinhardtii Pescadillo homolog (CHLREDRAFT_206018), partial
Description
Molecular Identification and Homology
CHLREDRAFT_206018 is annotated as a Pescadillo homolog in C. reinhardtii, sharing significant sequence similarity with Pescadillo proteins across eukaryotes . Key findings from BLAST analyses include:
| Species | Identity (%) | Positives (%) | E-value | Functional Domain Matches |
|---|---|---|---|---|
| Carya illinoinensis | 78.26 | 86.96 | 4.140e-17 | Ribosome assembly, nuclear localization |
| Medicago truncatula | 76.09 | 86.96 | 4.376e-17 | N-terminal β-propeller motifs |
| Arabidopsis thaliana | 65.22 | 78.26 | 2.890e-15 | C-terminal Yph1/Pescadillo motif |
| Oryza sativa | 78.95 | 86.84 | 7.984e-16 | rRNA processing activity |
Notably, residues 24–69 of the C. reinhardtii homolog align with conserved regions critical for interactions with ribosomal proteins and nucleolar localization signals .
Expression Systems in C. reinhardtii
The chloroplast of C. reinhardtii is a well-established platform for recombinant protein production due to:
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High transgene copy numbers (50–80 plastome copies per cell) .
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Post-translational folding support via endogenous chaperones (e.g., Spy) .
For CHLREDRAFT_206018, a codon-optimized sequence would likely be expressed under strong promoters like atpA or psbA, with epitope tags (e.g., HA) for detection . Typical yields for similar recombinant proteins in C. reinhardtii range from 6.77–22 mg/L in pilot-scale photobioreactors .
Stability and Downstream Processing
Key challenges in producing recombinant algal proteins include:
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Shear sensitivity: High-shear centrifugation (e.g., GEA Westfalia disc-stack) reduces cell integrity, increasing product loss to supernatants .
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Proteolytic degradation: Recombinant proteins show faster degradation rates (e.g., 50% Pal loss in 6–8 hours) compared to endogenous proteins like RbcL .
Optimized conditions for CHLREDRAFT_206018 would likely require:
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Reduced shear stress during harvest (Q/Σ < 65 × 10⁻⁹ m·s⁻¹) .
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Immediate post-harvest processing to minimize protease exposure .
Functional Implications
While direct functional data for the C. reinhardtii Pescadillo homolog is lacking, studies on orthologs suggest roles in:
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Ribosome maturation: Conserved Yph1/Pescadillo domains mediate 18S rRNA processing .
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Cell cycle control: Interaction with cyclin-dependent kinases in other species .
