Initial Metabolic Step of a Novel Ethanolamine Utilization Pathway and Its Regulation in <i>Streptomyces coelicolor</i> M145

Abstract

<i>Streptomyces coelicolor</i> is a Gram-positive soil bacterium with a high metabolic and adaptive potential that is able to utilize a variety of nitrogen sources. However, little is known about the utilization of the alternative nitrogen source ethanolamine. Our study revealed that <i>S. coelicolor</i> can utilize ethanolamine as a sole nitrogen or carbon (<i>N</i>/<i>C</i>) source, although it grows poorly on this nitrogen source due to the absence of a specific ethanolamine permease. Heterologous expression of a putative ethanolamine permease (SPRI_5940) from <i>Streptomyces</i><i>pristinaespiralis</i> positively influenced the biomass accumulation of the overexpression strain grown in defined medium with ethanolamine. In this study, we demonstrated that a glutamine synthetase-like protein, GlnA4 (SCO1613), is involved in the initial metabolic step of a novel ethanolamine utilization pathway in <i>S. coelicolor</i> M145. GlnA4 acts as a gamma-glutamylethanolamide synthetase. Transcriptional analysis revealed that expression of <i>glnA4</i> was induced by ethanolamine and repressed in the presence of ammonium. Regulation of <i>glnA4</i> is governed by the transcriptional repressor EpuRI (SCO1614). The <i>ΔglnA4</i> mutant strain was unable to grow on defined liquid Evans medium supplemented with ethanolamine. High-performance liquid chromatography (HPLC) analysis demonstrated that strain <i>ΔglnA4</i> is unable to utilize ethanolamine. GlnA4-catalyzed glutamylation of ethanolamine was confirmed in an enzymatic <i>in vitro</i> assay, and the GlnA4 reaction product, gamma-glutamylethanolamide, was detected by HPLC/electrospray ionization-mass spectrometry (HPLC/ESI-MS). In this work, the first step of ethanolamine utilization in <i>S. coelicolor</i> M145 was elucidated, and a putative ethanolamine utilization pathway was deduced based on the sequence similarity and genomic localization of homologous genes.<b>IMPORTANCE</b> Until now, knowledge of the utilization of ethanolamine in <i>Streptomyces</i> was limited. Our work represents the first attempt to reveal a novel ethanolamine utilization pathway in the actinobacterial model organism <i>S. coelicolor</i> through the characterization of the key enzyme gamma-glutamylethanolamide synthetase GlnA4, which is absolutely required for growth in the presence of ethanolamine. The novel ethanolamine utilization pathway is dissimilar to the currently known ethanolamine utilization pathway, which occurs in metabolome. The novel ethanolamine utilization pathway does not result in the production of toxic by-products (such as acetaldehyde); thus, it is not encapsulated. We believe that this contribution is a milestone in understanding the ecology of <i>Streptomyces</i> and the utilization of alternative nitrogen sources. Our report provides new insight into bacterial primary metabolism, which remains complex and partially unexplored.