Abstract
The obligate aerobic bacterium, Vitreoscilla, synthesizes elevated quantities of a homodimeric hemoglobin (VHb) under hypoxic growth conditions. Expression of VHb in heterologous hosts often enhances growth and product formation. A role in facilitating oxygen transfer to the respiratory membranes is one explanation of its cellular function. Immunogold labeling of VHb in both Vitreoscilla and recombinant Escherichia coli bearing the VHb gene clearly indicated that VHb has a cytoplasmic (not periplasmic) localization and is concentrated near the periphery of the cytosolic face of the cell membrane. OmpA signal-peptide VHb fusions were transported into the periplasm in E. coli, but this did not confer any additional growth advantage. The interaction of VHb with respiratory membranes was also studied. The K(d) values for the binding of VHb to Vitreoscilla and E. coli cell membranes were approximately 5-6 microm, a 4-8-fold higher affinity than those of horse myoglobin and hemoglobin for these same membranes. VHb stimulated the ubiquinol-1 oxidase activity of inverted Vitreoscilla membranes by 68%. The inclusion of Vitreoscilla cytochrome bo in proteoliposomes led to 2.4- and 6-fold increases in VHb binding affinity and binding site number, respectively, relative to control liposomes, suggesting a direct interaction between VHb and cytochrome bo.
Highlights
The obligate aerobic bacterium, Vitreoscilla, synthesizes elevated quantities of a homodimeric hemoglobin (VHb) under hypoxic growth conditions
This paper shows that the location of VHb in Vitreoscilla and recombinant E. coli is cytoplasmic and concentrated adjacent to the cell membrane, and evidence is provided that because of its membrane association properties, it may perform its cellular function by interacting directly with the respiratory apparatus of the cell
How the presence of VHb enables the bacterium to grow better under oxygen limitation is not conclusively known, but knowledge of its subcellular localization is vital to understanding its function
Summary
The obligate aerobic bacterium, Vitreoscilla, synthesizes elevated quantities of a homodimeric hemoglobin (VHb) under hypoxic growth conditions. The presence of a relatively large cellular concentration of VHb under oxygenlimiting conditions suggests that its primary function is to trap molecular oxygen and facilitate its transfer to the respiratory apparatus to enable Vitreoscilla to survive under these conditions despite its being a strict aerobe. It has been demonstrated through genetic engineering that the intracellular expression of VHb in various heterologous hosts often results in the enhancement of cell density, oxidative metabolism, engineered product formation, and bioremediation, especially under oxygen-limiting conditions.
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