Strzałka, Wojciech

Expression of three diadinoxanthin de-epoxidase genes of phaeodacylum tricornutum in escherichia coli origami b and BL21 strain by Monika Bojko( Book )

1 edition published in 2013 in English and held by 1 WorldCat member library worldwide

Violaxanthin conversion by recombinant diatom and plant de-epoxidases, expressed in escherichia coli : a comparative analysis by Monika Olchawa-Pajor( Book )

1 edition published in 2019 in English and held by 1 WorldCat member library worldwide

Heterologous expression of mlrA in a photoautotrophic host - Engineering cyanobacteria to degrade microcystins( )

1 edition published in 2018 in English and held by 1 WorldCat member library worldwide

Abstract: In this report, we establish proof-of-principle demonstrating for the first time genetic engineering of a photoautotrophic microorganism for bioremediation of naturally occurring cyanotoxins. In model cyanobacterium Synechocystis sp. PCC 6803 we have heterologously expressed Sphingopyxis sp. USTB-05 microcystinase (MlrA) bearing a 23 amino acid N-terminus secretion peptide from native Synechocystis sp. PCC 6803 PilA (sll1694). The resultant whole cell biocatalyst displayed about 3 times higher activity against microcystin-LR compared to a native MlrA host (Sphingomonas sp. ACM 3962), normalized for optical density. In addition, MlrA activity was found to be almost entirely located in the cyanobacterial cytosolic fraction, despite the presence of the secretion tag, with crude cellular extracts showing MlrA activity comparable to extracts from MlrA expressing E.coli . Furthermore, despite approximately 9.4-fold higher initial MlrA activity of a whole cell E.coli biocatalyst, utilization of a photoautotrophic chassis resulted in prolonged stability of MlrA activity when cultured under semi-natural conditions (using lake water), with the heterologous MlrA biocatalytic activity of the E.coli culture disappearing after 4 days, while the cyanobacterial host displayed activity (3% of initial activity) after 9 days. In addition, the cyanobacterial cell density was maintained over the duration of this experiment while the cell density of the E.coli culture rapidly declined. Lastly, failure to establish a stable cyanobacterial isolate expressing native MlrA (without the N-terminus tag) via the strong cpcB560 promoter draws attention to the use of peptide tags to positively modulate expression of potentially toxic proteins. Graphical abstract: Highlights: First description of heterologous microcystinase expression in a photoautotroph. MlrA overexpressing cyanobacteria shows higher activity than a natural gene host. Cyanobacteria chassis shows increased biomass stability in semi-natural system. Peptide tags may positively modulate expression of potentially toxic proteins. Abstract : Synechocystis sp. PCC 6803 overexpressing MlrA has shown about 3-fold higher whole-cell microcystin-detoxifying activity than natural MlrA gene host Sphingomonas sp. ACM 3962