Intracellular expression of Vitreoscilla hemoglobin modifies microaerobic Escherichia coli metabolism through elevated concentration and specific activity of cytochrome o.
The function of the reversible oxygen-binding hemoprotein from Vitreoscilla (VHb), which enhances oxygen-limited cell growth and recombinant protein production when functionally expressed in Escherichia coli, was investigated in wild-type E. coli and in E. coli mutants lacking one of the two terminal oxidases, cytochrome o complex (aerobic terminal oxidase, Cyo) or cytochrome d complex (microaerobic terminal oxidase, Cyd). Deconvolution of VHb, cytochrome o, and cytochrome d bands from in vivo absorption spectra revealed a 5-fold enhancement in cytochrome o content and a 1.5-fold increment in cytochrome d by VHb under microaerobic environments (dissolved oxygen less than 2% air saturation). Based upon oxygen uptake kinetics measurements of these mutants, the apparent oxygen affinity of the Cyo(+), Cyd(-) E. coli was increased in the presence of VHb, but no difference in the apparent K(m) was observed for the Cyo(-), Cyd(+) strain. Results suggest that the expression of VHb in E. coli increases the level and activity of terminal oxidases and thereby improves the efficiency of microaerobic respiration and growth.
Tsai
Philip S
PS
Institute of Biotechnology, ETH-Zürich, Switzerland.
Nägeli
Michael
M
Bailey
James E
JE
D000332:Aerobiosis; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D019149:Bioreactors; D019069:Cell Respiration; D002478:Cells, Cultured; D003001:Cloning, Molecular; D003573:Cytochrome b Group; D017937:Cytochrome d Group / Q000235:genetics / Q000378:metabolism; D003580:Cytochromes / Q000235:genetics / Q000378:metabolism*; D045222:Electron Transport Chain Complex Proteins; D003576:Electron Transport Complex IV / Q000378:metabolism*; D004734:Energy Metabolism; D004926:Escherichia coli / Q000145:classification / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D006454:Hemoglobins / Q000235:genetics / Q000378:metabolism*; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism; D010101:Oxygen Consumption; D011994:Recombinant Proteins / Q000378:metabolism; D012680:Sensitivity and Specificity; D013045:Species Specificity; D054793:Truncated Hemoglobins
D001426:Bacterial Proteins; D003573:Cytochrome b Group; D003580:Cytochromes; D045222:Electron Transport Chain Complex Proteins; D029968:Escherichia coli Proteins; D006454:Hemoglobins; D011994:Recombinant Proteins; D054793:Truncated Hemoglobins; C082613:hemoglobin protein, Vitreoscilla; D017937:Cytochrome d Group; C017168:cytochrome bo, E coli; D010088:Oxidoreductases; C040077:cytochrome bd terminal oxidase complex, E coli; C038862:cytochrome o oxidase; D003576:Electron Transport Complex IV; D010100:Oxygen