The algal chloroplast as a synthetic biology platform for production of therapeutic proteins

Abstract

The chloroplast of Chlamydomonas reinhardtii and other microalgae represents an attractive new platform for the synthesis of recombinant therapeutics using synthetic biology (synbio) approaches. Transgenes can be designed in silico, assembled from validated DNA parts and inserted at precise and predetermined locations within the chloroplast genome to give stable synthesis of a desired recombinant protein. Numerous recent examples of different therapeutic proteins produced successfully in the C. reinhardtii chloroplast highlight the potential of this green alga as a simple, low-cost and benign host. Furthermore, some of the features of the alga may offer additional advantages over more-established microbial, mammalian or plant-based systems. These include efficient folding and accumulation of the product in the chloroplast; a lack of contaminating toxins or infectious agents; reduced downstream processing requirements; the possibility to make complex therapeutics such as immunotoxins; and the opportunity to use the whole alga as a low-cost oral vaccine. In this paper we review the current status of algal chloroplast engineering with respect to therapeutic proteins. We also consider future advances in synbio tools, together with improvements to recipient strains, which will allow the design of bespoke strains with high levels of productivity.

Cyanobacterial Production of Biopharmaceutical and Biotherapeutic Proteins

“…Efforts to express human therapeutic proteins in photosynthetic microorganisms abound in the literature. In their preponderance, these entail heterologous transformation of microalgal chloroplasts as a synthetic biology platform for the production of biopharmaceutical and therapeutic proteins (Dyo and Purton, 2018, and references therein). The vast majority of such efforts have employed transformation of the chloroplast in the model green microalga Chlamydomonas reinhardtii via double homologous recombination of exogenous constructs encoding heterologous proteins (Demain and Vaishna, 2009;Surzycki et al, 2009;Tran et al, 2009;Coragliotti et al, 2011;Gregory et al, 2013;Jones and Mayfield, 2013;Rasala and Mayfield, 2015;Baier et al, 2018).…”

The Synthetic Biology Toolkit for Photosynthetic Microorganisms

“…Many genetic elements have been characterized in, or adapted to, microalgae. Several recent reviews have been published on algal synthetic biology and chloroplast genome modification (Doron et al, 2016;Scaife and Smith, 2016;Dyo and Purton, 2018;Poliner et al, 2018). Therefore, we will focus here on new developments and on nuclear genetic elements.…”

Multigenic engineering of the chloroplast genome in the green alga Chlamydomonas reinhardtii

“…DNA transformation of the algal chloroplast has been reported for a handful of species, but the technology is most advanced for C. reinhardtii [11]. Numerous studies have shown that insertion of foreign DNA into the plastome occurs exclusively via homologous recombination allowing precise and predictive targeting of transgenes into specific loci [12].…”

“…Our survey of the literature has revealed that over 100 different foreign proteins have been produced successfully in the C. reinhardtii chloroplast. These include a wide range of therapeutic proteins such as vaccines, hormones and antibodies [11]; industrial enzymes [15,16], and enzymes for synthesizing novel metabolites in the organelle [17][18][19][20][21]. However, in almost all cases the genetic engineering has involved the insertion of just one transgene (or a single transgene together with a bacterial gene such as aadA or aphA-6 as the selectable marker [13]).…”