The Functional Research Of Genes In Prokaryotic Ubiquitin-like Protein(Pup)-Proteasome System Of Mycobacterium

Objective: Pup-proteasome system is found in Mycobacterium smegmatis and Mycobacterium tuberculosis.It is used to control the degradation process of many kinds of proteins,such as misfolded proteins,regulatory proteins,toxic proteins,and so on.At the same time,it is related to pathogenicity,retention and amino acid circulation.The important components of Pup-proteasome system(Pup,Dop,proteasome)play a key role in the identification of target proteins and degradation.In this study,we investigated crucial genes encoding Pup-proteasome system,due to a number of key genes knockout,it will seriously affect and block Pup-proteasome system for normal operation,which is a clear understanding the role of the Pup-proteasome system in the physiological function of mycobacterium.By simulating a variety survival patterns,we can observe the dynamic changes of its occurrence and explore the relationship between Pup-proteasome system and Mycobacterium in a variety of stress conditions,in order to further explore the mechanism between Mycobacterium tuberculosis and immune system.Methods: The coding genes pup,dop,prcA and prcB were knocked out by homologous recombination,and the four genes were added into the knockout cells.The changes of growth rate of knockout and wild plants under normal conditions,anaerobic conditions,anaerobic dosing conditions and nutrient deficiency were detected.Results: Pup,dop,prcA,prcB of the four Pup-proteasome system related genes were successfully knocked out and recovered.In the normal culture condition,there was no significant change in the growth rate of the knockout plants,but the growth rate was slower in the later growth stage.The growth rate and the number of viable bacteria were decreased to some extent in the anaerobic stage.In addition,the growth rate of the knockout strain was significantly lower than that of the same anaerobic with isoniazid.In terms of nutrient deficiency,the growth rate of knockout plants decreased rapidly.Conclusion: The Pup-proteasome system is associated with the survival of Mycobacterium smegmatis under stress conditions.The results provide more clues for the Pup-proteasome system in Mycobacterium tuberculosis to maintain the persistence in host environment.It will build a foundation to explore the new mechanism of persistence of Mycobacterium tuberculosis.