Summary
Vitreoscilla hemoglobin is involved in oxygen metabolism of this bacterium, possibly in an unusual role for a microbe. We have isolated the Vitreoscilla hemoglobin structural gene from a pUC19 genomic library using mixed oligodeoxy-nucleotide probes based on the reported amino acid sequence of the protein. The gene is expressed in Escherichia coli from its natural promoter as a major cellular protein. The nucleotide sequence, which is in complete agrecment with the known amino acid sequence of the protein, suggests the existence of promoter and ribosome binding sites with a high degree of homology to consensus E. coli upstream sequences. In the case of at least some amino acids, a codon usage bias can be detected which is different from the biased codon usage pattern in E. coli. The down-stream sequence exhibits homology with the 3′ end sequences of several plant leghemoglobin genes. E. coli cells expressing the gene contain greater than fivefold more heme than controls.
Article PDF
Avoid common mistakes on your manuscript.
References
Boerman S, Webster DA (1982) Control of heme content in Vitreoscilla by oxygen. J. Gen Appl Microbiol 28:35–42
Dikshit KL, Webster DA (1987) Molecular cloning and restriction mapping of Vitreoscilla plasmid pVT1. Abstracts Annual meeting of American Society for Microbiology, Atlanta, Georgia, p 147
Glass RE (1982) Gene function-E. coli and its heritable elements. University of California Press, Berkeley, Los Angeles, pp 51–109
Gouy M, Gautier C (1982) Codon usage in bacteria: correlation with gene expressivity. Nucleic Acids Res 10:7055–7074
Hanahan D (1983) Studies on transformation of Escherichia coli with plasmids. J Mol Biol 166:557–580
Huynh TV, Young RA, Davis RW (1985) Construction and screening cDNA libraries in λgt10 and λgt11. In: Glover D (ed) DNA cloning: A practical approach. IRL Press, Oxford, pp 49–78
Iverson BL, Dervan PB (1987) Adenine specific DNA chemical sequencing, reaction. Nucleic Acids Res 15:7823–7830
Jarrell KF, Julseth C, Pearson B, Kuzio J (1987) Paucity of the Sau3AI recognition sequence (GATC) in the genome of Methanococcus voltae. Mol Gen Genet 208:191–194
Khosla C, Bailey JE (1988) Heterologous expression of a bacterial haemoglobin improves the growth properties of recombinant Escherichia coli. Nature 331:633–635
Kuhse J, Puehler A (1987) Conserved sequence motifs in the untranslated 3′ end of leghemoglobin transcripts isolated from broadbean nodules. Plant Sci 49:137–143
Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685
Lamba P, Webster DA (1980) Effect of growth conditions on yield and heme content of Vitreoscilla. J Bacteriol 142:169–173
Maniatis T, Fritsch EF, Sambrook J (1982) Molecular cloning — a laboratory manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York
Maxam AM, Gilbert W (1980) Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol 65:499–560
Merril CR, Goldman D, Sedman SA, Ebert MH (1983) Ultrasensitive stain for proteins in polyacrylamide gels shows regional variation in cerebrospinal fluid proteins. Science 211:1437–1438
Orii Y, Webster DA (1986) Photodissociation of oxygenated cytochrome o(s) (Vitreoscilla) and kinetic studies of reassociation. J Biol Chem 261:3544–3547
Silhavy TJ, Berman ML, Enquist LW (1984) Experiments with gene fusions Cold Spring Harbor Laboratory, Cold Spring Harbor, New York
Stougaard J, Sandal NN, Gron A, Kuehle A, Marcker KA (1987) 5′ analysis of the soyabean leghemoglobin lbc 3 gene: regulatory elements required for promoter activity and organ specificity. EMBO J 6:3565–3569
Tyree B, Webster DA (1978) The binding of cyanide and carbon monoxide to cytochrome o purified from Vitreoscilla. J Biol Chem 253:6988–6991
Wakabayashi S Matsubara H, Webster DA (1986) Primary sequence of a dimeric bacterial hemoglobin from Vitreoscilla. Nature 322:481–483
Webster DA, Liu CY (1974) Reduced nicotinamide adenine dinucleotide cytochrome o reductase associated with cytochrome o purified from Vitreoscilla. J Biol Chem 249:4257–4260
Author information
Authors and Affiliations
Additional information
Communicated by H. Hennecke
Rights and permissions
About this article
Cite this article
Khosla, C., Bailey, J.E. The Vitreoscilla hemoglobin gene: Molecular cloning, nucleotide sequence and genetic expression in Escherichia coli . Mol Gen Genet 214, 158–161 (1988). https://doi.org/10.1007/BF00340195
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF00340195