DOI: 10.4103/jpdtsm.jpdtsm_39_26 ISSN: 2949-6594

Codon-optimized Glucocerebrosidase cDNA Is Not Sufficient for Recombinant Expression in Pichia pastoris

Letícia Guerra Morosini, Patrícia Antonia Estima Abreu, Marcela Oliveira Vitarelli, Gisele Monteiro, Paulo Lee Ho
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BACKGROUND:

Gaucher disease is a lysosomal storage disorder caused by mutations in the GBA1 gene, leading to a deficiency of the enzyme glucocerebrosidase (GCase). Current enzyme replacement therapies are expensive due to production in mammalian systems. This study aimed to evaluate the expression of codon-optimized GCase in Pichia pastoris GlycoSwitch ® strains as a cost-effective alternative.

METHODS:

A codon-optimized GBA1 gene was cloned into the pJAG-s1 vector and used to transform P. pastoris GlycoSwitch ® colonies. Positive transformants were induced with methanol for 72 h. Protein expression was assessed in both supernatant and cell lysates using western blot analysis.

RESULTS:

Although successful cloning and transformation were confirmed by polymerase chain reaction and sequencing, no detectable GCase expression was observed in any of the analyzed colonies, either intracellularly or extracellularly.

CONCLUSIONS:

Codon optimization alone is insufficient to enable recombinant expression of GCase in P. pastoris . Additional strategies, including optimization of expression elements and posttranslational processing, are required.

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