(697e) Important Role of Class I Heat Shock Genes on the Butyrate Production and the Response to Low pH Condition By Clostridium Tyrobutyricum ATCC 25755

Class I heat shock genes (HSGs) code for molecular chaperones which play a major role in the bacterial response to sudden increases of environmental stress by assisting protein folding. The objective of this study was to identify the expression profile of HSGs of Clostridium tyrobutyricum under butyric acid stress, and explore the effect of overexpressing these genes on butyric acid production. The qRT-PCR analysis shows that five Class I HSPs genes of the C. tyrobutyricum significantly up-regulated in response to butyric acid or butyrate. At a butyric acid concentration of 6g/L, the expressions of grpE, DnaK, DnaJ, groESL, htpG increased 2.3, 1.8, 0.7, 2.7, 2.6, 1.0- fold. Meanwhile, the expressions of those genes increased 6.2, 8.6, 4.7, 8.9, 9.2, 2.6-fold at a butyrate concentration of 10g/L, respectively. Overexpression of DnaK, groESL, htpG significantly improved the tolerance to butyric acid. At a butyric acid concentration of 10g/L, the groESL-overexpressing strains grew faster, and can achieve higher cell density (OD₆₀₀ 2.0 vs OD₆₀₀1.4 for the wild type). Under butyrate stress, the groESL- and htpG-overexpressing strains had higher inhibition rate constant (Kp=3.52g/L, Kp=2.56 vs Kp=1.75g/L for the wild type), suggesting that both of them have higher tolerance to butyrate. The free-cell fermentation results show that groESL-overexpressing strains stopped producing butyric acid at a butyrate concentration of 44.25±2.60 g/L, which was higher compared to the wild type (34.52±1.92 g/L). The results showed that metabolic engineering to improve the butyric acid tolerance and production has a potential application prospect.