Improving astaxanthin production in Escherichia coli by co-utilizing CrtZ enzymes with different substrate preference

Background The bifunctional enzyme β-carotene hydroxylase (CrtZ) catalyzes the hydroxylation of carotenoid β-ionone rings at the 3, 3’ position regardless of the presence of keto group at 4, 4’ position, which is an important step in the synthesis of astaxanthin. The level and substrate preference of CrtZ may have great effect on the amount of astaxanthin and the accumulation of intermediates. Results In this study, the substrate preference of PC crtZ from Paracoccus sp. PC1 and PA crtZ from Pantoea Agglomerans were certified and were combined utilization for increase astaxanthin production. Firstly, PC crtZ from Paracoccus sp. PC1 and PA crtZ from P. Agglomerans were expressed in platform strains CAR032 (β-carotene producing strain) and Can004 (canthaxanthin producing strain) separately to identify their substrate preference for carotenoids with keto groups at 4,4’ position or not. The results showed that PC crtZ led to a lower zeaxanthin yield in CAR032 compared to that of PA crtZ . On the contrary, higher astaxanthin production was obtained in Can004 by PCcrtZ than that of PA crtZ . This demonstrated that PCCrtZ has higher canthaxanthin to astaxanthin conversion ability than PACrtZ, while PACrtZ prefer using β-carotene as substrate. Finally, Ast010, which has two copies of PA crtZ and one copy of PC crtZ produced 1.82 g/L of astaxanthin after 70 h of fed-batch fermentation. Conclusions Combined utilization of crtZ genes, which have β-carotene and canthaxanthin substrate preference respectively, can greatly enhance the production of astaxanthin and increase the ratio of astaxanthin among total carotenoids.