Study On The Function Of NrgA In Paenibacillus Polymyxa SC2-M1 Based On Proteomics And Metabolomics

NrgA is a high-affinity ammonium transporter located on the cell membrane.It is conserved in all areas of life.It is responsible for the transport of ammonium ions on the membrane,and only works when the concentration of ammonium ions is low.The mutation of NrgA gene will cause its ammonium ion transport ability to be weakened or lost,so the study believes that NrgA is indispensable for maintaining the steady state of the intracellular environment.Previous proteomics data from the laboratory showed that in Paenibacillus polymyxa SC2-M1,the NrgA gene may be involved in antibiotic synthesis.In this study,homologous recombination was used to knock out the NrgA gene in Paenibacillus polymyxa SC2-M1,combined with proteomics and metabolomics to further explore the function of NrgA in Paenibacillus polymyxa.Compared with the wild-type SC2-M1,the ammonium ion transport capacity of the mutant strain was significantly weakened,the flagella and its motility were lost,and the performance of antagonistic bacteria was enhanced.The differentially expressed proteins of wild strain SC2-M1 and mutant strain NrgA-q were detected by 4D LFQ quantitative proteomics.A total of 20 differentially expressed proteins were identified,13 of which were up-regulated and 7 were down-regulated.The functional classification of the differentially expressed proteins revealed that the differentially expressed proteins were mainly concentrated in the transport and metabolism of various substances,including the transport and metabolism of amino acids,nucleotides,carbohydrates,coenzymes,and lipids.Enrichment analysis of differential protein metabolism pathways found that NrgA is closely related to purine metabolism pathway and spermidine / ornithine metabolism pathway,NrgA may change the expression levels of purF,iunH3,ade,potD3 and potA11 by affecting the transport of ammonium ions,thereby affecting purine metabolism and spermidine / ornithine transport.Combined with the qPCR results of fluorescence and analysis of flagellar traits,it was found that NrgA may affect the expression of cheW9 throughthe transport of ammonium ions and thus affect the generation and motility of the flagella of the strain.Using non-target metabolomics to detect the differential metabolites of wild strain SC2-M1 and mutant strain NrgA-q,a total of 242 differential metabolites were identified,127 were up-regulated and 115 were down-regulated.There are 73 differential metabolites.Enrichment analysis of differential metabolite pathways found that NrgA is likely to affect purine metabolism pathways through ammonium ion transport.Combined with the analysis of the plate bacterial antagonism test,it was found that NrgA may affect the ability of the strain to antagonize the bacteria by affecting the synthesis or metabolism of Sedecamycin.Through joint analysis of proteomics and metabolomics,it was found that NrgA protein is one of the important ammonium ion transport pathways at the whole cell level in Paenibacillus polymyxa,and its absence obviously affects the purine metabolism in cell basic metabolism.This study found a variety of functions of NrgA in Paenibacillus polymyxa,including affecting the expression of cheW9 to affect the flagella generation and motility of the strain;affecting the synthesis or metabolism of Sedecamycin to affect the ability of the strain to antagonize the bacteria;affecting purine metabolism Routes,etc.,have important theoretical value for us to understand the transport of ammonium ions in Paenibacillus polymyxa.