Identification And Functional Study Of Spermidine Synthase SpeD In Lysobacter Enzymogenes OH11

Lysobacter enzymogenes belongs to the genus of Lysobacter of Xanthomodaceae family.Lysobacter enzymogenes OH 11 is emerging as a novel biocontrol agent,which was isolated from cayenne-root soil in laboratory.This bacterial genomic can produce a variety of secondary metabolic antibacterial substances,such as chitinase,protease cellulase,?-1,3-glucanas and Heat-Stable Antifungal Factor-HSAF,which with high DNA G+C%,no flagella but have gliding ability.HSAF(Heat-Stable Antifungal Factor)produced by Lysobacter enzymogenes exhibits a new mode of action.Polyamines are low molecular weight aliphatic nitrogen containing two or more amino residues,it exists in animal,plants,fungi and bacteria,such as putrescine,spermine and spermidine.However,using further bioinformatics study,we found that speD,which codes a spermidine synthase,is a close neighbor of the clp gene.This speD-clp pair is further found to be widely distributed in many bacterial genomes,such as Pseudomonas aeruginosa and Xanthomonas oryzae,suggesting their co-conservation may imply important functions together.Here,using HPLC,we showed that SpeD is active in catalyzing spermidine synthesis in L.enzymogenes OH11.Genetic complementarity and in vitro experiments showed that,similar to Clp,SpeD is also required for HSAF production,Twitiching Motility and resitiant to kinds of pathogens,as evidenced by the fact that the deficiency of the speD mutant in producing HSAF can be rescued by the exogenous addition of spermidine,which establishes a solid link between the enzymatic activitve of SpeD and its regulatory effect.Also,by using Western Blot,SpeD can stabilize the expression of Clp on translation level,but not transcription level.Importantly,in vitro experiments,I also provide the first example that SpeD,but not spermidine,directly interacts with its neighbor transcription regulation Clp.In this study,we find firstly that a polyamine biosynthesis enzyme SpeD can interact with a transcription regulator Clp.Furthermore,exogenous addition of spermidine seem to stabilize Clp protein,and spermidine inhabitior DCHA can cancel it,resulting in a coordinated regulation of HSAF biosynthesis by the SpeD-Clp protein pair,which provides a new cognitive for the follow-up scientistfic workers.