Study On Fermentation Character And Proteomic Qualification On A Glutamine-producing Corynebacterium Glutamicum G32

Glutamine,an important and useful amino acid,not only has important physiology significance,but also is a prospective new medicine.Corynebacterium glutamicum G32 could excessively accumulate glutamine when the ammonium concentration is high while the production of glutamate decreases respectively.This result demonstrated that the nitrogen metabolism and the metabolic regulation of this mutant are trememdously different from the standard strain Corynebacterium glutamicum 13032.We tried to figure it out the response of C.glutamicum G32 to diffrent ammonium concentrations utilizing proteome analysis in this paper.Glutamate dehydrogenase(GDH),glutamine synthetase(GS),glutamate synthase(GOGAT) has been considered as the major ammonia assimilatory enzymes in Corynebacterium glutamicum.Characteristics of the three enzymes in different nitrogen source were examined.The results showed that the all three enzymes frome the cells in medium at high nitrogen level were enhanced and change of pH had no effect on the specific activities of the three enzymes.The activity of glutamate synthase changed greatly in diferrent nitrogen source.It showed that glutamate dehydrogenase and glutamate synthase were the important enzymes in nitrogen metabolism.GDH is the key enzyme,which is related to carbon metabolism with nitrogen metabolism,and it is very flexile.Compared to C.glutamicum G32 growing in medium at low nitrogen level,the metabolism of bacteria inoculated in medium at high nitrogen level apparently turned to another direction:the utility of glucose fastened,PPP process enhanced,TCA circulation before acetone dicarboxylic acid strengthened,but enzymes of TCA after acetone carboxylic acid act differently,some of them enhanced while some weakened. The proteins related to the growth of cells are dramatically lower.At the end of fermentation,the proteins function to degrade free oxgen were enhanced,which imply that it's oxygen free radical that damge the cells to stop the fermentation.Three key genes glnA,glnA2 and amtR were cloned and sequenced.The results showed that only the 47th amino acid of the glutamine synthatase I in C.glutamicum G32 change from Ile to Val,which couldn't change the physiological function of it.There are two gaps happened in glA2 and it probably to change its function while it also has the possibility to weaken the transcription of another regulatory gene glnE. The central regulatory factor AmtR has four amino-acid changes which could influence the whole nitrogen regulatory mechanism of this bacterium.