Immobilization and stability of a Rhizopus oryzae lipase expressed in Pichia pastoris: Comparison between native and recombinant variants

Abstract

The stability of a soluble extract containing a recombinant lipase from Rhizopus oryzae (Cursive) lipase (rROL) produced by Pichia pastoris (Cursive), as well as that for the commercial extract containing the lipase produced by the native organism (nROL), was investigated. The results showed higher residual activity values of the commercial protein compared with the recombinant one. Moreover, two different kinds of support, the polypropylene powder EP100 and Eupergit®C, were tested to immobilize the enzymes. The residual activity of the immobilizated derivatives was also tested to determine whether their stability was enhanced. The results showed a slight improvement in rROL using both supports but a decrease in nROL using Eupergit®C. The study of the residual activity of soluble and immobilized enzymes was performed by means of a central composite rotatable experiment design. In addition, EP100 adsorption isotherms were determined.

Truncated Prosequence of Rhizopus oryzae Lipase: Key Factor for Production Improvement and Biocatalyst Stability

“…As can be seen in Figure 7B, proROL lost only 10% of its activity after 24 h of incubation at 25 • C and pH 8.12. Therefore, the temperature had a stronger effect than pH on proROL stability, which is consistent with the results of previous studies on rROL stability [5].…”

“…Because proROL has been deemed more stable than the enzyme consisting of the mature sequence only [11], proROL stability was compared to rROL stability under three different conditions selected from a previously done experimental design carried out for rROL characterization [5]. Such conditions were chosen on the grounds that they previously led to a variable loss of activity.…”

“…3.1.1.3) are hydrolytic enzymes that catalyze the hydrolysis of esters in aqueous media. Also, they are widely known for carrying out synthesis, interesterification, and, especially significant for the biodiesel industry, transesterification reactions in organic media [5]. However, two of the most significant restrictions to the industrial use of lipases as biocatalysts in biodiesel production are their costs and their poor stability, notably in the presence of alcohols, such as methanol [6], which limits their reusability and compromises the economic feasibility of the bioprocess.…”

Fast and economic immobilization methods described for non-commercial Pseudomonaslipases

“…Supports were pretreated following an adaptation from Guillen and coworkers [37] where the polypropylene matrices were activated with ethanol (3 mL per gram of MP1000 or EP100), followed by alcohol removal by vacuum filtration. Immobilization was carried out by mixing the pretreated supports with 5 mL cell extract or supernatant samples, following a ratio of 1 mg protein per g of EP100 and 2 mg protein per g of MP1000.…”

Towards optimal substrate feeding for heterologous protein production in Pichia pastoris (Komagataella spp) fed‐batch processes under PAOX1 control: a modeling aided approach

“…Thus, experimental comparison with the most suitable alternatives previously reported was carried out after the definition and analysis of the optimized profiles (OP) by numerical methods using a process model for systems with substrate inhibition patterns on both q P and μ . As a model protein, 1–3 positional selective recombinant Rhizopus oryzae lipase (ROL) was selected because of its multiple applications in pharma, food and energy industries …”