Abstract
The ribosome and tRNA levels of Escherichia coli cells, transformed with a native or mutated Vitreoscilla hemoglobin genes (vhb), were investigated using asymmetrical flow field-flow fractionation (AFFFF). Mutagenesis of vhb by error-prone PCR was carried out to alter the growth behavior of microaerobically cultivated native VHb-expressing E. coli. A VHb mutant, pVMT1, was identified, which was able to reach a remarkably high final A600 of 15, the value of which being 160% higher than that of a VHb control carrying pVHb8 (A600 5.8). AFFFF revealed that cells expressing mutant vhbs showed up to a doubling in the number of active 70S ribosomes cell−1, an almost 3-fold increase in the number of tRNAs cell−1, and up to a 26% increase in the mass fraction of active 70S ribosomes.
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Andersson CIJ, Holmberg N, Farrés J, Bailey JE, Bülow L, Kallio PT (2000) Error-prone PCR of Vitreoscilla hemoglobin (VHb) to support the growth of microaerobic Escherichia coli. Biotechnol. Bioeng. 70: 446–455.
Bashford D, Chotia C, Lesk AM (1987) Determinants of a protein fold. Unique features of the globin amino acid sequences. J. Mol. Biol. 196: 199–216.
Chen R, Bailey JE (1994) Energetic effect of Vitreoscilla hemoglobin expression in Escherichia coli: an on-line P31-NMR and saturation transfer study. Biotechnol. Prog. 10: 360–364.
Holmberg N, Lilius G, Bailey JE, Bülow L (1997) Transgenic tobacco expressing Vitreoscilla hemoglobin exhibits enhanced growth and altered metabolite production. Nat. Biotechnol. 15: 244–247.
Kallio PT, Kim DJ, Tsai PS, Bailey JE (1994) Intracellular expression of Vitreoscilla hemoglobin alters Escherichia coli energy metabolism under oxygen-limited conditions. Eur. J. Biochem. 219: 201–208.
Khosla C, Bailey JE (1988) Heterologous expression of a bacterial haemoglobin improves the growth properties of recombinant Escherichia coli. Nature 331: 633–635.
Litzén A (1993) Separation speed, retention and dispersion in asymmetrical flow field-flow fractionation as functions of channel dimensions and flow rates. Anal. Chem. 65: 461–470.
Nilsson M, Birnbaum S, Wahlund KG (1996) Determination of relative amounts of ribosome and subunits in Escherichia coli using asymmetrical flow field-flow fractionation. J. Biochem. Biophys. Meth. 33: 9–23.
Nilsson M, Bülow L, Wahlund KG (1997) Use of flow field-flow fractionation for the rapid quantitation of ribosome and ribosomal subunits in Escherichia coli at different protein production conditions. Biotechnol. Bioeng. 54: 461–467.
Nilsson M, Kallio PT, Bailey JE, Bülow L, Wahlund KG (1999) Expression of Vitreoscilla hemoglobin in Escherichia coli enhances ribosome and tRNA levels: a flow field-flow fractionation study. Biotechnol. Prog. 15: 158–163.
Spirin AS, Belitsina NV, Lishnevskaya EB (1972) On some artifacts of sucrose gradient sedimentation of ribosomes. FEBS Lett. 24: 219–224.
Tarricone C, Galizzi A, Coda A, Ascenzi P, Bolognesi M (1997) Unusual structure of the oxygen-binding site in the dimeric bacterial hemoglobin from Vitreoscilla sp. Structure 5: 497–507.
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Andersson, C.I., Arfvidsson, C., Kallio, P.T. et al. Enhanced ribosome and tRNA contents in Escherichia coli expressing a truncated Vitreoscilla hemoglobin mutant analyzed by flow field-flow fractionation. Biotechnology Letters 25, 1499–1504 (2003). https://doi.org/10.1023/A:1025475703006
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DOI: https://doi.org/10.1023/A:1025475703006