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Subcellular distribution of enzymes involved in α-glucan utilization in Klebsiella pneumoniae

Proteins of cytoplasm, periplasm, cytoplasmic and outer membrane

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Abstract

The double-isotopic labelling technique was used to identify comprehensively proteins involved in α-glucan catabolism in Klebsiella pneumoniae NCTC 9633. Cells were grown with either glycerol in the presence of 3H-leucine or with glycerol plus maltose in the presence of 14C-leucine. Each labelled culture was then fractionated into the main subcellular components, i.e. the cytoplasm, periplasm, cytoplasmic and outer membrane. Corresponding fractions derived from 3H-labelled and 14C-labelled cells were combined, and the proteins were analyzed by polyacrylamide gel electrophoresis under denaturing conditions. Gel slices were then counted for 3H- and 14C-radioactivity, a positive deviation from the standard 14C/3H ratio being evidence for the presence of a protein specifically induced by maltose in the culture medium. The protein pattern thus obtained was compared with the properties of proteins comprising a similar pathway for maltodextrin utilization in Escherichia coli K-12. Ample information which has been obtained mainly by genetic analysis is available about maltodextrin-utilizing enzymes in E. coli K-12.

  1. 1.

    Cytoplasm. Neither amylomaltase nor maltodextrin phosphorylase, well-known soluble enzymes, were identifiable by the double-labelling technique, presumably because these enzymes constitute only a very minor portion of all soluble proteins in the cytoplasm.

  2. 2.

    Periplasm. A prominent protein with a mass of 43000 daltons (43 kD) was found similar to the maltose-binding protein of E. coli K-12 (44 kD).

  3. 3.

    Cytoplasmic membrane. At least 2 proteins with a mass between 40 and 50 kD were detected, minor proteins were seen at ≈ 15 and ≈ 20 kD. One or 2 of the proteins may function as a permease catalyzing the active transport of maltodextrins.

  4. 4.

    Outer membrane. The major protein had a mass of 55 kD, other proteins were found with ≈ 18, ≈48, and ≈140 kD. The major protein may have the same function as the maltodextrin pore protein in E. coli K-12 (55 kD), because K. pneumoniae could grow on 10 μM maltose at practically the same rate as on 10 mM maltose. The 140 kD protein is pullulanase.

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References

  • Buchanan, R. E., Gibbons, N. E. (eds.): Bergey's manual of determinative bacteriology. Baltimore, Md.: Williams and Wilkins 1974

    Google Scholar 

  • Essig, U.: Amylomaltase aus Klebsiella pneumoniae 1009/III. Diplomarbeit, Univ. Freiburg (1976)

  • Fenzl, F., Decker, M., Haas, D., Tanner, W.: Characterization and partial purification of an inducible protein related to hexose proton cotransport in Chlorella vulgaris. Europ. J. Biochem. 72, 509–514 (1977)

    Google Scholar 

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

    Google Scholar 

  • Hazelbauer, G. L.: Maltose chemoreceptor of Escherichia coli. J. Bacteriol. 122, 206–214 (1975a)

    Google Scholar 

  • Hazelbauer, G. L.: Role of the receptor for bacteriophage lambda in the functioning of the maltose chemoreceptor of Escherichia coli. J. Bacteriol. 124, 119–126 (1975b)

    Google Scholar 

  • Heppel, L. A., Harkness, D. R., Hilmoe, R. J.: A study of the substrate specificity and other properties of the alkaline phosphatase of Escherichia coli. J. Biol. Chem. 237, 841–846 (1962)

    Google Scholar 

  • Hofnung, M.: Divergent operons and the genetic structure of the maltose B region in Escherichia coli K-12. Genetics 76, 169–184 (1974)

    Google Scholar 

  • Hofnung, M., Hatfield, D., Schwartz, M.: mal B Region in Escherichia coli K-12: Characterization of new mutations. J. Bacteriol. 117, 40–47 (1974)

    Google Scholar 

  • Hofnung, M., Jezierska, A., Braun-Breton, C.: lam B Mutations in E. coli K-12: Growth of host range mutants and effect of nonsense suppressors. Molec. Gen. Genet. 145, 207–213 (1976)

    Google Scholar 

  • Kellermann, O., Szmelcman, S.: Active transport of maltose in Escherichia coli K-12. Involvement of a “periplasmic” maltose binding protein. Europ. J. Biochem. 47, 139–149 (1974)

    Google Scholar 

  • Kiehn, E. D., Holland, J. J.: Membrane and nonmembrane proteins of mammalian cells. Organ, species and tumor specificities. Biochemistry 9, 1729–1738 (1970)

    Google Scholar 

  • Kolber, A. D., Stein, W. D.: Identification of a component of a transport “carrier” system: Isolation of the permease expression of the lac operon of Escherichia coli. Nature (London) 209, 691–694 (1966)

    Google Scholar 

  • Linder, D., Kurz, G., Bender, H., Walldenfels, K.: 1,4-α-Glucan phosphorylase from Klebsiella pneumoniae. Purification, subunit structure and amino acid composition. Europ. J. Biochem. 70, 291–303 (1976)

    Google Scholar 

  • Löhr, G. W., Waller, H. D.: Glucose-6-phosphat-Dehydrogenase. In: Methoden der enzymatischen Analyse (H. U. Bergmeyer, ed.), pp. 673–681 Weinheim: Verlag Chemie 1974

    Google Scholar 

  • Lowry, D. H., Rosebrough, N. J., Farr, A. L., Randall, R.: Protein measurement with the Folin phenol reagent. J. Biol. Chem. 193, 265–275 (1951)

    Google Scholar 

  • Mizushima, S., Yamada, H.: Isolation and characterization of two outer membrane preparations from Escherichia coli. Biochim. Biophys. Acta 375, 44–53 (1975)

    Google Scholar 

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

    Google Scholar 

  • Osborn, M. J., Gander, J. E., Parisi, E., Carson, J.: Mechanism of assembly of the outer membrane of Salmonella typhimurium. J. Biol. Chem. 247, 3962–3972 (1972)

    Google Scholar 

  • Palm, D., Thanner, F., Schwenk, L.: Purification and properties of Escherichia coli maltodextrin phosphorylase. In: Metabolic interconversion of enzymes 1975 (S. Shaltiel, ed.), pp. 37–43. Berlin-Heidelberg-New York: Springer 1976

    Google Scholar 

  • Palmer, T. N., Ryman, B. E., Whelan, W. J.: The action pattern of amylomaltase from Escherichia coli. Europ. J. Biochem. 69, 105–115 (1976)

    Google Scholar 

  • Palmer, T. N., Wöber, G., Whelan, W. J.: The pathway of exogenous and endogenous carbohydrate utilization in Escherichia coli: a dual function for the enzymes of the maltose operon. Europ. J. Biochem. 39, 601–612 (1973)

    Google Scholar 

  • Randall-Hazelbauer, L. L., Schwartz, M.: Isolation of the bacteriophage lambda receptor from Escherichia coli K-12. J. Bacteriol. 116, 1436–1446 (1973)

    Google Scholar 

  • Schwartz, M.: Espression phénotypique et localisation génétique de mutations affectant le métabolisme du maltose chez Escherichia coli K-12. Ann. Inst.Pasteur 112, 673–700 (1967)

    Google Scholar 

  • Schwartz, M., le Minor, L.: Occurrence of the bacteriophage lambda receptor in some Enterobacteriaceae. J. Virol. 15, 679–685 (1975)

    Google Scholar 

  • Szmelcman, S., Hofnung, M.: Maltose transport in Escherichia coli K-12: Involvement of the bacteriophage, lambda receptor. J. Bacteriol. 124, 112–118 (1975)

    Google Scholar 

  • Wöber, G.: Pullulanase is a characteristic of many Klebsiella species and functions in the degradation of starch. Europ. J. Appl. Microbiol. 3, 71–80 (1976)

    Google Scholar 

  • Wöhner, G., Wöber, G.: Pullulanase, an enzyme of starch catabolism, is associated with the outer membrane of Klebsiella. Arch. Microbiol. 116, 303–310 (1978)

    Google Scholar 

  • Wolf-Watz, H., Normark, S., Bloom, G. D.: Rapid method for isolation of large quantities of outer membrane from Escherichia coli K-12 and its application to the study of envelope mutants. J. Bacteriol. 115, 1191–1197 (1973)

    Google Scholar 

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  1. Biochemie (Fachbereich Chemie) der Philipps-Universität, Lahnberge, D-3550, Marburg/Lahn 1, Germany

    Gerhard Wöhner & Günter Wöber

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  1. Gerhard Wöhner
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  2. Günter Wöber
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Wöhner, G., Wöber, G. Subcellular distribution of enzymes involved in α-glucan utilization in Klebsiella pneumoniae . Arch. Microbiol. 116, 311–316 (1978). https://doi.org/10.1007/BF00417857

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