The Expression of NOX From Synthetic Promoters Reveals an Important Role of the Redox Status in Regulating Secondary Metabolism of Saccharopolyspora erythraea.

Xiaobo Li,Peter R Jensen,Ju Chu

doi:10.3389/fbioe.2020.00818

Abstract

Redox cofactors play a pivotal role in primary cellular metabolism, whereas the clear link between redox status and secondary metabolism is still vague. In this study we investigated effects of redox perturbation on the production of erythromycin in Saccharopolyspora erythraea by expressing the water-forming NADH oxidase (NOX) from Streptococcus pneumonia at different levels with synthetic promoters. The expression of NOX reduced the intracellular [NADH]/[NAD+] ratio significantly in S. erythraea which resulted in an increased production of erythromycin by 19∼29% and this increment rose to 60% as more oxygen was supplied. In contrast, the lower redox ratio resulted in a decreased production of another secondary metabolite, the reddish pigment 7-O-rahmnosyl flaviolin. The metabolic shifts of secondary metabolism results in a higher NADH availability which compensates for its oxidization via NOX. The expression of the erythromycin biosynthesis gene cluster (BGC) in the NOX-expression strains was upregulated as the activity of diguanylate cyclase was inhibited moderately by NADH. This study also suggested that lower intracellular [NADH]/[NAD+] ratio benefits the biosynthesis of erythromycin by potentially affecting the biosynthesis of the secondary messenger, bis-(3′-5′)-cyclic dimeric guanosine monophosphate (c-di-GMP), which may stimulate the positive regulation of erythromycin BGC via BldD. The present work provides a basis for future cofactor manipulation in S. erythraea to improve the industrial production of erythromycin.

Highlights

Derivatives of currently prevalent macrolides, e.g., erythromycin, are among the most frequently used chemotherapeutic agents for treating infections
It has been proven previously that the nucleotide-based secondary messenger, bis(3 -5 )-cyclic dimeric guanosine monophosphate (c-di-GMP), even at very low concentrations activates the binding between BldD and the biosynthesis gene cluster (BGC) (Tschowri et al, 2014; Xu et al, 2019). c-diGMP allows a rapid integration of external and internal signals into fine-tuned regulatory pathways, which controls the cellular responses to changing environmental conditions (Tschowri et al, 2014) but the signals involved in the c-di-GMP-related regulatory networks are still unknown
The objective of the present work is to explore the possibility of improving erythromycin production by fine tuning the intracellular redox status in S. erythraea and elucidate the signals and mechanisms triggering the physiological response toward disturbed NADH/NAD+ ratio

Summary

Introduction

Derivatives of currently prevalent macrolides, e.g., erythromycin, are among the most frequently used chemotherapeutic agents for treating infections. The biosynthesis of erythromycin is involved in a complex secondary metabolic network, that is connected with the primary metabolism via specific precursors and cofactors, including a supply of NADPH as reducing equivalents (McPherson et al, 1998). The biosynthesis of erythromycin is linked to signal transduction and transcriptional regulation. The transcription of the erythromycin biosynthesis gene cluster (BGC) is under the positive regulation by BldD, which can bind all the promoter regions in the BGC (Chng et al, 2008). It has been proven previously that the nucleotide-based secondary messenger, bis(3 -5 )-cyclic dimeric guanosine monophosphate (c-di-GMP), even at very low concentrations activates the binding between BldD and the BGC (Tschowri et al, 2014; Xu et al, 2019). It has been proven previously that the nucleotide-based secondary messenger, bis(3 -5 )-cyclic dimeric guanosine monophosphate (c-di-GMP), even at very low concentrations activates the binding between BldD and the BGC (Tschowri et al, 2014; Xu et al, 2019). c-diGMP allows a rapid integration of external and internal signals into fine-tuned regulatory pathways, which controls the cellular responses to changing environmental conditions (Tschowri et al, 2014) but the signals involved in the c-di-GMP-related regulatory networks are still unknown

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Conclusion