As a novel protein post-translational modification, lysine succinylation is widely involved in metabolism regulation. In this study, we focused on the distribution of lysine succinylation sites and their physiological functions in Saccharopolyspora erythraea. Using high-resolution 4D label free mass spectrometry, a large and global protein succinylome was identified in a hypersuccinylated strain E3ΔsucC. The results showed that succinylated proteins are predominantly involved in protein synthesis-related pathways (e.g., ribosomes, tRNA) and metabolic pathways, such as the TCA cycle. Proteins in these pathways generally have a higher lysine content, suggesting that lysine succinylation may have a greater regulatory role in biochemical reactions involving acidic substrates. Motif analysis revealed that charged amino acids (D, E, K, R and W) display a more regular distribution around acylation sites, implying that the polar effect between residues may be the key factor influencing lysine succinylation. Based on predicted protein structures, we highlighted the potential impact of lysine succinylation on enzyme activity in the TCA cycle. In conclusion, this study offers valuable insights into the regulation of lysine succinylation and contributes to a comprehensive understanding of its physiological functions in actinomyces.

Omics approaches have been applied to understand the boosted productivity of natural products by industrial high-producing microorganisms. Here, with the updated genome sequence and transcriptomic profiles derived from high-throughput sequencing, we exploited comparative omics analysis to further enhance the biosynthesis of erythromycin in an industrial overproducer, Saccharopolyspora erythraea HL3168 E3. By comparing the genome of E3 with the wild type NRRL23338, we identified fragment deletions inside 56 coding sequences and 255 single nucleotide polymorphisms over the genome of E3. Substantial numbers of genomic variations were observed in genes responsible for pathways which were interconnected to the biosynthesis of erythromycin by supplying precursors/cofactors or by signal transduction. Through comparative transcriptomic analysis, L-glutamine/L-glutamate and 2-oxoglutarate were identified as reporter metabolites. Around the node of 2-oxoglutarate, genomic mutations were also observed. Furthermore, the transcriptomic data suggested that genes involved in the biosynthesis of erythromycin were significantly up-regulated constantly, whereas some genes in biosynthesis clusters of other secondary metabolites contained nonsense mutations and were expressed at extremely low levels. Based on the omics association analysis, readily available strategies were proposed to engineer E3 by simultaneously overexpressing sucB (coding for 2-oxoglutarate dehydrogenase E2 component) and sucA (coding for 2-oxoglutarate dehydrogenase E1 component), which increased the erythromycin titer by 71% compared to E3 in batch culture. This work provides more promising molecular targets to engineer for enhanced production of erythromycin by the overproducer.