Vitreoscilla hemoglobin - Intracellular localization and binding to membranes

The obligate aerobic bacterium, Vitreoscilla, synthesizes elevated quantiti es of a homodimeric hemoglobin (VHb) under hypoxic growth conditions, Expre ssion of VHb in heterologous hosts often enhances growth and product format ion. A role in facilitating oxygen transfer to the respiratory membranes is one explanation of its cellular function. Immunogold labeling of VHb in bo th Vitreoscilla and recombinant Escherichia coli bearing the VHb gene clear ly indicated that VHb has a cytoplasmic (not periplasmic) localization and is concentrated near the periphery of the cytosolic face of the cell membra ne. OmpA signal-peptide VHb fusions were transported into the periplasm in E. coli, but this did not confer any additional growth advantage. The inter action of VHb with respiratory membranes was also studied. The K-d values f or the binding of VHb to Vitreoscilla and E. coli cell membranes were simil ar to5-6 muM, a 4-8-fold higher affinity than those of horse myoglobin and hemoglobin for these same membranes. VHb stimulated the ubiquinol-l oxidase activity of inverted Vitreoscilla membranes by 68%. The inclusion of Vitre oscilla cytochrome bo in proteoliposomes led to 2.4- and 8-fold increases i n VHb binding affinity and binding site number, respectively, relative to c ontrol liposomes, suggesting a direct interaction between VHb and cytochrom e bo.