| Streptococcus thermophilus is widely used in the production of fermented dairy products as an important fermentative strain,and high density culture is one of the most important technical links in the preparation of Streptococcus thermophilus.Streptococcus thermophilus belongs to amino acid auxotrophic strain so that amino acid metabolism plays an important role in resisting the stress of adverse environmental conditions and the growth of Streptococcus thermophilus.In terms of the high density culture of Streptococcus thermophilus,it is particularly important to study the nutritional requirements and metabolic mechanism in specific proliferative environments,especially amino acid metabolism as a source of nitrogen.The deficiency of this regard may has certain occasionality and blindness for the study on screening of nitrogen sources,and meanwhile it may impede the precise design of high density nitrogen sources and the availability of high biomass.This paper targeted the growth dynamics of Streptococcus thermophilus TF96 with good fermentation characteristics cultivated in chemical synthetic medium,which using amino acid as the sole nitrogen source under the control of pH.With the application of high-throughput transcriptional Sequencing and metabolomics,the change of key genes transcription levels and metabolite levels in the amino acid metabolic pathways were studied,in order to indicate which key genes involved in the metabolic regulation of amino acids and which metabolites produced in this regulatory mode during the proliferation of Streptococcus thermophilus TF96.The results of transcriptome analysis showed that amino acid metabolism was one of the most active metabolic pathways of Streptococcus thermophilus TF96,which the genes involved in amino acid metabolism accounted for 15.37% of the total number of genes.Among the transcripts related to amino acid metabolism,the genes involved in tyrosine metabolism were the most,followed by glycine,serine,threonine,arginine,proline,cysteine and methionine.The genes involved in tyrosine metabolism accounted for 15% of the total amino acid metabolic genes,14% for glycine,serine and threonine,also for arginine and proline,about 12% for cysteine and methionine.The key genes involved in the regulation of alanine metabolism were ala A,alr,dat,ddl and dltA.The metabolites of alanine were poly-D-propionyl-propionyl and D-alanyl-D-alanine under the control of the above key genes.Based on transcriptomics and metabolomics data,the metabolic flow of alanine was mainly the synthesis of peptidoglycan and phosphate acid.The key genes involved in the regulation of valine,leucine and isoleucine metabolism were ilvA,tdcB,ilvB,ilvG,ilvI and ilvE,and the intermediates of isopropyl malic acid were obtained under the control of the above key genes.Based on transcriptomics and metabolomics data,valine,leucine and isoleucine were significantly higher in T3,and it was founded that branched chain amino acids were used to the proliferation of Streptococcus thermophilus TF96,and the metabolic flow of branched chain amino acids were amino acids synthesis pathways.The key genes involved in the regulation of aspartate metabolism were argH,aspB,nadB and lysC.The metabolites of aspartic acid were 5-hydroxytetrahydropyrimidine,L-4-aspartyl-P and N6-acetyl-LL-2,6-diaminopimelic acid,etc.Based on transcriptomics and metabolomics data,the metabolic flow of aspartic acid was the tetrahydropyrimidine synthesis pathway and lysine synthesis pathway in order to regulate the osmotic pressure and stabilize the protein structure of the cells.The key genes involved in the regulation of glutamic acid,arginine and proline metabolism were glnA,gdhA,carB,gadA,glmS,OTC,arcC,argH and amiE.Under the control of the above key genes,the metabolites were D-glutamic acid,5-oxo-D-proline,N-acetylglutamic acid,N2-succinyl-ornithine,?-feruloylagmatine and 1-pyrroline-4-hydroxy-2-carboxylate etc.Based on transcriptomics and metabolomics data,the metabolic flow of glutamate was mainly 5-oxo-D-proline synthesis pathway and ornithine synthesis pathway,and arginine was N2-succinyl-ornithine and ?-Feruloylate pathway,and proline was aminopentanoic acid and 1-pyrroline-4-hydroxy-2-carboxylate pathways.The key genes involved in the regulation of histidine metabolism were hisB and METTL6.The metabolites of histidine were hercynine and ergothioneine.Based on transcriptomics and metabolomics data,the metabolic flow of histidine was predominantly betaine synthesis pathway to exert the cytoprotective effect of antioxidant and osmotic pressure.The key genes involved in the regulation of cysteine and methionine metabolism were cysE,cysK,metK,gadA,metE,mtnN,aspB and metC.The metabolites were L-cystine,L-homocysteine,Sadenosyl-L-homocysteine,S-ribose-L-homocysteine,L-methionine-oxide and taurine,etc.Based on transcriptomics and metabolomics data,the metabolic flow of cysteine and methionine was mainly methionine synthesis pathway and taurine pathway.The key genes involved in the regulation of serine,glycine and threonine metabolism were DLD,lpd,gly A,tdcG,sdaA,sdaB and soxA.The metabolites were O-acetyl-serine,cysteine,O-succinylserine and cystathionine.Based on transcriptomics and metabolomics data,the metabolic flow was mainly the cysteine synthesis pathway.The key genes involved in the regulation of phenylalanine metabolism were dat,dadA and aspB.The metabolites were phenylacetaldehyde,phenylacetic acid,benzoic acid,D-phenylalanine,cinnamic acid and vanillin,etc.Based on transcriptomics and metabolomics data,the metabolic flow was mainly phenylacetic acid pathway,phenylpyruvic acid pathway and cinnamic acid pathway.The key genes involved in the regulation of tyrosine metabolism were adhE,aspB and METTL6.The metabolites were L-DOPA,dopamine,noradrenaline,adrenaline,3,4-dihydroxy-phenyllactate,dopaquinone,melanin,tyramine,4-hydroxyphenylacetaldehyde,4-hydroxyphenylactate,homoprotocatechuate,hydroxyl-phenylpyruvate and homogentisate,etc.Based on transcriptomics and metabolomics data,the metabolic flow of tyrosine was mainly catecholamine pathway,melanin pathway and oxidative decomposition pathway.The key genes involved in the regulation of tryptophan metabolism were amiE,and the metabolites were indole,hydroxyindole,N-hydroxytryptamine,indole-3-acetaldehyde,indole ethanol,indoleacetate,2-formaminobenzoylacetate,anthranilate,5-hydroxy-L-tryptophan,serotonin,5-methyloxyindoleacetate,5-hydroxy-N-formylkynurenine,etc.Based on transcriptomics and metabolomics data,the metabolic flow of tryptophan was predominantly serotonin pathway and kynurenine pathway.The key genes involved in the regulation of glutathione metabolism were gor,pepD,G6 PD and zwf.The metabolites of were L-?-glutamylcysteine,glutathione disulfide,glutathionyl-spermidine,glutathionyl-spermine and glutathionyl-aminopropyl-cadaverine.Based on transcriptomics and metabolomics data,the metabolic flow of glutathione was mainly glutathionyl-spermidine,glutathionyl-spermine and glutamylcysteine pathway.The key genes involved in the regulation of selenium compounds were metE,and the metabolites were selenocystathionine,selenohomocysteine,selenomethionine,Se-methyl-selenocysteine and ?-glutamyl-se-methylselenocysteine.Based on transcriptomics and metabolomics data,the metabolic flow of selenium compounds was mainly methyl-selenomethionine and methylselenocysteine synthesis pathway.Amino acid metabolism pathway of Streptococcus thermophilus TF96 could greatly affect the proliferation and growth of bacterial cells.It could be seen that the original concentration of arginine,serine,cysteine,methionine,aspartic acid and glutamic acid could be appropriately increased in the medium,and the concentration of histidine was appropriately increased in the mid-exponential growth phase.Meanwhile L-alanine in the original medium was replaced with D-alanine.In contrast,the original concentrations of proline,phenylalanine,tyrosine and tryptophan were appropriately reduced,both to avoid the occurrence of nitrogen wastage and reduce the inhibition of cell proliferation. |
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