Abstract
Enzyme immobilization to solid matrices often presents a challenge due to protein conformation sensitivity, desired enzyme purity, and requirements for the particular carrier properties and immobilization technique. Surface display of enzymes at the cell walls of microorganisms presents an alternative that has been the focus of many research groups worldwide in different fields, such as biotechnology, energetics, pharmacology, medicine, and food technology. The range of systems by which a heterologous protein can be displayed at the cell surface allows the appropriate one to be found for almost every case. However, the efficiency of display systems is still quite low. The most frequently used yeast for the surface display of proteins is Saccharomyces cerevisiae. However, apart from its many advantages, Saccharomyces cerevisiae has some disadvantages, such as low robustness in industrial applications, hyperglycosylation of some heterologous proteins, and relatively low efficiency of surface display. Thus, in the recent years the display systems for alternative yeast hosts with better performances including Pichia pastoris, Hansenula polymorpha, Blastobotrys adeninivorans, Yarrowia lipolytica, Kluyveromyces marxianus, and others have been developed. Different strategies of surface display aimed to increase the amount of displayed protein, including new anchoring systems and new yeast hosts are reviewed in this paper.
Highlights
Recombinant protein expression and incorporation into the yeast cell wall represents an exceptional tool for engineering and displaying many heterologous proteins
In the last two decades, much effort has been focused on the development of molecular tools for surface display in different microorganisms, in yeasts
Most attempts aimed to improve the applicability of surface display was directed at increasing in the amount of heterologous protein imbedded at the surface of the cell
Summary
Recombinant protein expression and incorporation into the yeast cell wall represents an exceptional tool for engineering and displaying many heterologous proteins. To improve the efficiency and enable surface anchoring of protein complexes, new expression systems and novel techniques for the display of heterologous proteins have been developed. Surface display systems employ genes encoding heterologous proteins fused with the complete fragments of genes coding forcell yeast wall[1,2,3,4]. The most frequently used yeast for surface display is Saccharomyces cerevisiae and its cell wall proteins α-agglutinin, a-agglutinin, and Flo. Numerous cell-surface display systems in which the N-terminus the heterologous protein is fused to the Flo flocculation domain were constructed. Display efficiency of immobilized proteins can be improved by deletion of endogenous PIR genes, and in the case of two PIR proteins fused with the enzyme of interest are co-expressed simultaneously in the yeast cell [1]. Many attempts to increase protein display efficiency in S. cerevisiae have been undertaken including optimization of cell wall anchoring domains, transcription and translation levels of heterologous proteins [38,39], as well as post-translational modifications and transfer in secretory pathway that might limit successfulness of heterologous protein display
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