Abstract:
:Respiratory NADH oxidation in the rumen bacterium Prevotella bryantii is catalyzed by the Na+-translocating NADH:quinone oxidoreductase (NQR). A method for cell disruption and membrane isolation of P. bryantii under anoxic conditions using the EmulisFlex-C3 homogenizer is described. We compared NQR activity and protein yield after oxic and anoxic cell disruption by the EmulsiFlex, by ultrasonication, and by glass beads treatment. With an overall membrane protein yield of 50 mg L-1 culture and a NADH oxidation activity of 0.8 µmol min-1 mg-1, the EmulsiFlex was the most efficient method. Anoxic preparation yielded fourfold higher NQR activity compared to oxic preparation. P. bryantii lacks genes coding for superoxide dismutases and cell extracts do not exhibit superoxide dismutase activity. We propose that inactivation of NQR during oxic cell rupture is caused by superoxide, which accumulates in P. bryantii extracts exposed to air. Anoxic cell rupture is indispensable for the preparation of redox-active proteins and enzymes such as NQR from P. bryantii.
journal_name
Arch Microbioljournal_title
Archives of microbiologyauthors
Schleicher L,Fritz G,Seifert J,Steuber Jdoi
10.1007/s00203-019-01805-xsubject
Has Abstractpub_date
2020-07-01 00:00:00pages
1263-1266issue
5eissn
0302-8933issn
1432-072Xpii
10.1007/s00203-019-01805-xjournal_volume
202pub_type
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