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Cloning of mglB, the structural gene for the galactose-binding protein of Salmonella typhimurium and Escherichia coli

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Summary

From libraries of EcoRI fragments of Salmonella thyphimurium and Escherichia coli DNA in λgt7, phages could be isolated that carry mglB, the structural gene of the galactose-binding protein as well as other mgl genes. Lysogenization of an E. coli mutant carrying a defective galactose-binding protein with λgt7 mglB (Salmonella) restores full galactose transport and galactose chemotaxis. Both the E. coli mutant protein as well as the wild-type Salmonella galactose-binding protein are synthesized in this strain. The EcoR1 fragments of both organisms carrying the mgl genes were 6 Kb long. They were subcloned into the multicopy plasmid pACUC184. The hybrid plasmid containing the Salmonella mgl DNA gives rise to the synthesis of large amounts of galactose-binding protein in the periplasm of E. coli. The protein can be precipitated by antibodies against the E. coli binding protein and is identical to the fully processed protein isolated from Salmonella typhimurium LT2. In vitro protein synthesis (Zubay-system) with either λgt7 mgl phages as well as the hybrid plasmid as DNA matrix produces the galactose-binding protein mainly in precursor form that is precipitable by specific antibodies.

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References

  • Anraku Y (1968) Transport of sugars and amino acids in bacteria. J Bill Chem 243:3116–3128

    Google Scholar 

  • Ardeshir F, Higgins CF, Ferro-Luzzi Ames G (1981) Physical map of the Salmonella typhimurium histidine transport operon: Correlation with the genetic map. J Bacteriol 147:401–409

    Google Scholar 

  • Birnboim HC, Doly J (1979) A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucl Acid Res 7:1513–1523

    Google Scholar 

  • Boos W (1972) Structurally defective galactose-binding protein isolated from a mutant negative in the β-methylgalactoside transport system of Escherichia coli. J Biol Chem 247:5414–5424

    Google Scholar 

  • Boos W (1974) Pro and contra carrier proteins; sugar transport via the periplasmic galactose-binding protein. Current Top Membr Transp 5:51–136

    Google Scholar 

  • Boos W, Gordon AS, Hall RE, Price HR (1972) Transport properties of the galactose-binding protein of Escherichia coli; substrate induced conformational change. J Biol Chem 247:917–924

    Google Scholar 

  • Boos W, Steinacher I, Engelhardt-Altendorf D (1981) Mapping of mglB, the structural gene of the galactose-binding protein of Escherichia coli. Mol Gen Genet, submitted

  • Chang ACY, Cohen SN (1978) Construction and characterization of amplifiable multicopy DNA cloning vehicles derived from the P15A cryptic miniplasmid. J Bacteriol 134:1141–1156

    Google Scholar 

  • Davis RW, Botstein D, Roth JR (1980) Advanced Bacterial Genetics. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY

    Google Scholar 

  • Hatfield D, Hofnung M, Schwartz M (1969) Genetic analysis of the maltose A region in Escherichia coli. J Bacteriol 98:559–567

    Google Scholar 

  • Hazelbauer GL, Adler J (1971) Role of the galactose-binding protein in the chemotaxis of Escherichia coli toward galactose. Nature New Biol 12:101–104

    Google Scholar 

  • Inouye M, Halegouya S (1980) Secretion and membrane localization of proteins in Escherichia coli. CRC Crit Rev Biochem 7:339–371

    Google Scholar 

  • Johnson WC, Silhavy TJ, Boos W (1975) Two-dimensional polyacrylamide gel electrophoresis of envelope proteins of Escherichia coli. Appl Microbiol 29:405–413

    Google Scholar 

  • Kessler SW (1975) Rapid isolation of antigens from cells with a staphylococcal protein A-antibody adsorbent: Parameters of the interaction of antibody-antigen complexes with protein. A. J Immunol 115:1617–1624

    Google Scholar 

  • Laemmli UK (1970) Cleavage of structural protein during the assembly of the head of bacteriophage T4. Nature (London) 227:680–685

    Google Scholar 

  • Lederberg EM, Cohen SN (1974) Transformation of Salmonella typhimurium by plasmid desoxyribonucleic acid. J Bacteriol 119:1072–1074

    Google Scholar 

  • Lengeler J, Hermann KO, Unsöld HJ, Boos W (1971) The Regulation of the β-methylgalactoside transport system and of the galactose-binding protein of Escherichia coli K12. Eur J Biochem 19:457–470

    Google Scholar 

  • Mahoney WC, Hogg RW, Hermodson MA (1981) The amino acid sequence of the D-galactose-binding protein from Escherichia coli B/r. J Biol Chem 256:4350–4356

    Google Scholar 

  • McGowan EB, Silhavy TJ, Boos W (1974) Involvement of a tryptophan residue in the binding site of Escherichia coli galactose-binding protein. Biochemistry 13:993–999

    Google Scholar 

  • Miller DM III, Olson JS, Quiocho FA (1980) The mechanism of sugar binding to the periplasmic receptor for galactose chemotaxis and transport in Escherichia coli. J Biol Chem 255:2465–2471

    Google Scholar 

  • Miller JH (1972) Experiments in molecular genetics. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY

    Google Scholar 

  • Neu HC, Heppel LA (1965) The release of enzymes from Escherichia coli by osmotic shock and during the formation of spheroplasts. J Biol Chem 240:3685–3692

    Google Scholar 

  • Ordal GW, Adler J (1974) Isolation and complementation of mutants in galactose taxis and transport. J Bacteriol 117:509–516

    Google Scholar 

  • Parnes JR, Boos W (1973) Unidirectional transport activity mediated by the galactose-binding protein of Escherichia coli. J Biol Chem 248:4436–4445

    Google Scholar 

  • Quiocho FA, Plugrath JW (1980) The structure of D-galactose-binding protein at 4.1 Å resolution looks like arabinose-binding protein. J Biol Chem 255:6559–6561

    Google Scholar 

  • Randall LL, Hardy SJS (1977) Synthesis of exported proteins by membrane-bound polysomes from Escherichia coli. Eur J Biochem 75:43–53

    Google Scholar 

  • Schumacher G, Bussmann K (1978) Cell-free synthesis of proteins related to sn-glycerol-3-phosphate transport in Escherichia coli. J Bacteriol 135:239–250

    Google Scholar 

  • Schumann W, Bade EG (1979) In vitro constructed plasmids containing both ends of bacteriophage Mu DNA express phage functions. Mol Gen Genet 169:97–105

    Google Scholar 

  • Shen BHP, Boos W (1973) Regulation of the β-methylgalactoside transport system and the galactose-binding protein by the cell cycle of Escherichia coli. Proc Natl Acad Sci USA 70:1481–1485

    Google Scholar 

  • Shuman HA, Silhavy TJ, Beckwith JR (1980) Labeling of proteins with β-galactosidase by gene fusion. J Biol Chem 255:168–174

    Google Scholar 

  • Silhavy TJ, Szmelcman S, Boos W, Schwartz M (1975) On the significance of the retention of ligand by protein. Proc Natl Acad Sci USA 72:2120–2124

    Google Scholar 

  • Silhavy TJ, Hartig-Beecken I, Boos W (1976) Periplasmic protein related to the sn-glycerol-3-phosphate transport system of Escherichia coli. J Bacteriol 126:951–958

    Google Scholar 

  • Smith WP, Tai PC, Thompson RC, Davis BD (1977) Extracellular labeling of nascent polypeptides traversing the membrane of Escherichia coli. Proc Natl Acad Sci USA 74:2830–2834

    Google Scholar 

  • Thomas M, Cameron JR, Davis RW (1974) Viable molecular hybrids of bacteriophage lambda and eukaryotic DNA. Proc Natl Acad Sci USA 71:4579–4583

    Google Scholar 

  • Zubay G (1973) In vitro synthesis of protein in microbial systems. Annu Rev Genet 7:267–287

    Google Scholar 

  • Zukin RS, Strange PC, Heavy LR, Koshland DE Jr (1977) Properties of the galactose-binding protein of Salmonella typhimurium and Escherichia coli. Biochemistry 16:381–386

    Google Scholar 

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Communicated by E. Bautz

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Müller, N., Heine, HG. & Boos, W. Cloning of mglB, the structural gene for the galactose-binding protein of Salmonella typhimurium and Escherichia coli . Mol Gen Genet 185, 473–480 (1982). https://doi.org/10.1007/BF00334143

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  • DOI: https://doi.org/10.1007/BF00334143

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