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In Silico Cloning of a Phe-free \(\kappa\)-casein in Kluyveromyces lactis  using cheese whey as c-source for the production of cream cheese for Phenylketonuria patients
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  • Paola Sanabria Ramírez,
  • Martha Diaz,
  • Laura Buitrago,
  • Maria Alejandra Escobar Holguin,
  • Miguel Menco,
  • Javier Lopez
Paola Sanabria Ramírez
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Martha Diaz
University of Los Andes (Uniandes)
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Laura Buitrago
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Maria Alejandra Escobar Holguin
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Miguel Menco
University of Los Andes (Uniandes)
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Javier Lopez
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Abstract

Phenylketonuria (PKU) is a genetic disorder that increases the levels of Phenylalanine (Phe) in the blood due to a defect in the gene that helps create the enzyme necessary to break down this amino acid in the uptake of several protein products. In consequence, PKU patients have limited access to suitable protein sources to complement their daily diets.  In this research, it is proposed the in silico cloning of a Phe-free k-casein as an alternative source of protein for PKU patients, and its use in a cream cheese formulation. The cloning process is proposed in K. lactis as hos microorganism using cheese whey as c-source and an expression vector built with from pKLAC2 integrated with a Phe-free \(\kappa\)-casein gene codon-optimized for its expression by secretion in K. lactis. Additionally, it is proposed a downstream separation process combining a centrifugation unit for the biomass separation, and filtration and ultrafiltration units for the Phe-free \(\kappa\)-casein purification. Finally, it was proposed a formulation for Phe-free cream cheese, including Phe-free k-casein (12% wt.), canola oil (14% wt.), lactase (3% wt.), salt (7% wt.) and water (csp).