High yield recombinant penicillin G amidase production and export into the growth medium using Bacillus megaterium.
BACKGROUND
During the last years B. megaterium was continuously developed as production host for the secretion of proteins into the growth medium. Here, recombinant production and export of B. megaterium ATCC14945 penicillin G amidase (PGA) which is used in the reverse synthesis of beta-lactam antibiotics were systematically improved.


RESULTS
For this purpose, the PGA leader peptide was replaced by the B. megaterium LipA counterpart. A production strain deficient in the extracellular protease NprM and in xylose utilization to prevent gene inducer deprivation was constructed and employed. A buffered mineral medium containing calcium ions and defined amino acid supplements for optimal PGA production was developed in microscale cultivations and scaled up to a 2 Liter bioreactor. Productivities of up to 40 mg PGA per L growth medium were reached.


CONCLUSIONS
The combination of genetic and medium optimization led to an overall 7-fold improvement of PGA production and export in B. megaterium. The exclusion of certain amino acids from the minimal medium led for the first time to higher volumetric PGA activities than obtained for complex medium cultivations.
Yang
Yang
Y
Biochemical Engineering, TU-BCE, HZI-Helmholtz Centre for Infection Research, Inhoffenstrasse 7, 38124 Braunschweig, Germany. yya@helmholtz-hzi.de
Biedendieck
Rebekka
R
Wang
Wei
W
Gamer
Martin
M
Malten
Marco
M
Jahn
Dieter
D
Deckwer
Wolf-Dieter
WD