Efficient itaconic acid production from glycerol with Ustilago vetiveriae TZ1

Thiemo Zambanini, Hamed Hosseinpour Tehrani, Elena Geiser, Dorothee Merker, Sarah Schleese, Judith Krabbe, Joerg M. Buescher, Guido Meurer, Nick Wierckx, Lars M. Blank

The family of Ustilaginaceae is known for their capability to naturally produce industrially valuable chemicals from different carbon sources. Recently, several Ustilaginaceae were reported to produce organic acids from glycerol, which is the main side stream in biodiesel production. In this study, we present Ustilago vetiveriae as new production organism for itaconate synthesis from glycerol. In a screening of 126 Ustilaginaceae, this organism reached one of the highest titers for itaconate combined with a high-glycerol uptake rate. By adaptive laboratory evolution, the production characteristics of this strain could be improved. Further medium optimization with the best single colony, U. vetiveriae TZ1, in 24-deep well plates resulted in a maximal itaconate titer of 34.7 ± 2.5 g L(-1) produced at a rate of 0.09 ± 0.01 g L(-1) h(-1) from 196 g L(-1) glycerol. Simultaneously, this strain produced 46.2 ± 1.4 g L(-1) malate at a rate of 0.12 ± 0.00 g L(-1) h(-1). Due to product inhibition, the itaconate titer in NaOH-titrated bioreactor cultivations was lower (24 g L(-1)). Notably, an acidic pH value of 5.5 resulted in decreased itaconate production, however, completely abolishing malate production. Overexpression of ria1 or mtt1, encoding a transcriptional regulator and mitochondrial transporter, respectively, from the itaconate cluster of U. maydis resulted in a 2.0-fold (ria1) and 1.5-fold (mtt1) higher itaconate titer in comparison to the wild-type strain, simultaneously reducing malate production by 75 and 41%, respectively. The observed production properties of U. vetiveriae TZ1 make this strain a promising candidate for microbial itaconate production. The outcome of the overexpression experiments, which resulted in reduced malate production in favor of an increased itaconate titer, clearly strengthens its potential for industrial itaconate production from glycerol as major side stream of biodiesel production.