Enhancement of productivity of recombinant alpha-amidating enzyme by low temperature culture

We have produced a recombinant C-terminal alpha-amidating enzyme (799BglII alpha-AE) derived from Xenopus laevis by culturing a CHO cell line named 3 mu-1S. Recently, we demonstrated that culturing 3 mu-1S cells at a temperat ure below 37 degrees C led to the following phenomena: inhibited cell growt h with high viability, enhanced cellular productivity (maximally at 32 degr ees C), and suppressed medium consumption and release of impurities from th e cells. Therefore, it is suggested that the 799BglII alpha-AE production w ill be increased by culturing a sufficient number of the cells at a low tem perature (especially at 32 degrees C). To assess this effect on batch and p erfusion cultures, the culture temperature was shifted from 37 to 32 degree s C in the mid-exponential phase in the case of batch culture and from 37 t o 34 degrees C when the cell density became high enough in the case of perf usion culture. Application of the low temperature culture to batch and perf usion cultures was effective in comparison with the culture at 37 degrees C : the productivity per medium and the productivity per time were increased severalfold with enhanced cellular productivity at a low culture temperatur e. The low temperature culture also increased the relative content of 799Bg lII alpha-AE in the supernatant and reduced the glucose consumption. The me thod presented here would contribute to production of bioactive proteins us ing other recombinant cell lines.