An aqueous two-phase system to pre-purify a heterologously produced siderophore

Abstract

Natural products span broad activities and applications; however, their access and production are often limited by native cellular sources. As a result, the heterologous production of a siderophore termed yersiniabactin (Ybt) was completed using the surrogate host Escherichia coli. Post-production and purifi cation steps are complicated by the complex nature of most media used for cell growth, prompting the development in this work of an aqueous two-phase pre-purifi cation system capable of rapidly and simply enhancing the concentration of the target Ybt compound. INNOVATIONTh e biological sources of natural products result in production processes that are complex in media composition and complicated compound purifi cation. Such is the case for the heterologous production of yersiniabactin (Ybt) using E. coli. Production through E. coli establishes an engineering-friendly platform for compound access and eliminates the need to handle the native producer Yersinia pestis, a priority pathogen. In addition, the fi nal compound has several potential applications in metal sequestration. However, these and other options will benefi t from an economical (rapid and requiring simple reagents), green (solvent-free) and simple (non-chromatographic) pre-purifi cation step. Toward this end, we employed an aqueous two-phase system (ATPS) for the enhanced separation of Ybt from a crude cellular extract. Th e separation approach is unique in its application to Ybt and enables a straightforward means of simplifying purifi cation strategies for future applications in metal removal, retrieval and reuse. Th e approach also off ers broader potential as an option for other natural products produced in heterologous formats.