caa a22 4500 606212264 CHVBK 20210128101042.0 cr unu---uuuuu 210128e20150701xx s 000 0 eng 10.1007/s10295-015-1623-8 doi (NATIONALLICENCE)springer-10.1007/s10295-015-1623-8 Growth of Bacillus methanolicus in 2M methanol at 50°C: the effect of high methanol concentration on gene regulation of enzymes involved in formaldehyde detoxification by the ribulose monophosphate pathway [Elektronische Daten] [Ahmet Bozdag, Claire Komives, Michael Flickinger] Bacillus methanolicus MGA3 is a Gram-positive aerobic methylotroph growing optimally at 50-53°C. Methylotrophy in B. methanolicus is encoded on pBM19 and by two chromosomal copies of the methanol dehydrogenase (mdh), hexulose phosphate synthase (hps) and phosphohexuloisomerase (phi) genes. However, there are no published studies on the regulation of methylotrophy or the dominant mechanism of detoxification of intracellular formaldehyde in response to high methanol concentration. The µ max of B. methanolicus MGA3 was assessed on methanol, mannitol and glucose. B. methanolicus achieved a µ max at 25mM initial methanol of 0.65±0.007h−1, which decreased to 0.231±0.004h−1 at 2M initial methanol. Slow growth was also observed with initial methanol concentrations of >2M. The µ max on mannitol and glucose are 0.532±0.002 and 0.336±0.003h−1, respectively. Spiking cultures with additional methanol (100mM) did not disturb the growth rate of methanol-grown cells, whereas, a 50mM methanol spike halted the growth in mannitol. Surprisingly, growth in methanol was inhibited by 1mM formaldehyde, while mannitol-grown cells tolerated 2mM. Moreover, mannitol-grown cells removed formaldehyde faster than methanol-grown cells. Further, we show that methanol oxidation in B. methanolicus MGA3 is mainly carried out by the pBM19-encoded mdh. Formaldehyde and formate addition down-regulate the mdh and hps genes in methanol-grown cells. Similarly, they down-regulate mdh genes in mannitol-grown cells, but up-regulate hps. Phosphofructokinase (pfk) is up-regulated in both methanol and mannitol-grown cells, which suggests that pfk may be a possible synthetic methylotrophy target to reduce formaldehyde growth toxicity at high methanol concentrations. Society for Industrial Microbiology and Biotechnology, 2015 Methylotrophy nationallicence Bacillus methanolicus nationallicence Formaldehyde tolerance nationallicence Formaldehyde toxicity nationallicence RUMP pathway nationallicence Bozdag Ahmet Department of Plant and Microbial Biology, North Carolina State University, Raleigh, NC, USA aut Komives Claire Department of Chemical Engineering, San Jose State University, San Jose, CA, USA aut Flickinger Michael Golden LEAF Biomanufacturing Training and Education Center, North Carolina State University, Campus Box 7928, 27695-7928, Raleigh, NC, USA aut Journal of Industrial Microbiology & Biotechnology Springer Berlin Heidelberg 42/7(2015-07-01), 1027-1038 1367-5435 42:7<1027 2015 42 10295 https://doi.org/10.1007/s10295-015-1623-8 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s10295-015-1623-8 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Bozdag Ahmet Department of Plant and Microbial Biology, North Carolina State University, Raleigh, NC, USA aut NATIONALLICENCE 700 1- Komives Claire Department of Chemical Engineering, San Jose State University, San Jose, CA, USA aut NATIONALLICENCE 700 1- Flickinger Michael Golden LEAF Biomanufacturing Training and Education Center, North Carolina State University, Campus Box 7928, 27695-7928, Raleigh, NC, USA aut NATIONALLICENCE 773 0- Journal of Industrial Microbiology & Biotechnology Springer Berlin Heidelberg 42/7(2015-07-01), 1027-1038 1367-5435 42:7<1027 2015 42 10295