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Chapter 7 : Cell Fractionation

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Abstract:

This chapter presents techniques for the fractionation of cellular components, including organelles and appendages, beginning with external surfaces and ending with internal components. It should be appreciated, however, that many of the techniques described in the chapter, from cell breakage to centrifugation theory, will have similar applications. The factors pertinent to cell fractionation, often dictating the approach needed, are the differences in wall structures and the bonding forces within walls (and associated surface materials) responsible for maintaining cell shape. Purification of the murein-outer membrane complex from the crude envelope fraction by density gradient separation in sucrose is described. Lipopolysaccharide (LPS) isolated by the phenol-water method preserves the reactivity of pseudomonad LPS with antibodies in Western immunoblots and is better in this regard than LPS isolated by the Darveau-Hancock method. In general, the murein portion of the cell wall is first removed to form protoplasts or spheroplasts, which are then lysed to provide either the plasma membrane or the plasma plus outer membranes. A structurally intact, intracytoplasmic membrane array may best be isolated by lysis of osmotically sensitive spheroplasts. The effect of lysis and physiological conditions on the nucleoid is discussed by Korch.

Citation: Koval S, Sprott G. 2007. Cell Fractionation, p 108-137. In Reddy C, Beveridge T, Breznak J, Marzluf G, Schmidt T, Snyder L (ed), Methods for General and Molecular Microbiology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817497.ch7

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Bacteria and Archaea
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Viruses
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Chemicals
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Plasma Membrane
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Outer Membrane Proteins
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FIGURE 1

Isolation of cytoplasmic membranes by discontinuous sucrose gradient centrifugation. A membrane fraction from spheroplast lysate in 33% sucrose was loaded on 40 to 50% sucrose steps and centrifuged at 4°C for 48 h (110,000 × , max) in a Beckman SW27 swinging-bucket rotor. A thin section of the upper band revealed closed vesicles bounded by the double-track membrane, typical of cytoplasmic membranes. See reference 135 for further details.

Citation: Koval S, Sprott G. 2007. Cell Fractionation, p 108-137. In Reddy C, Beveridge T, Breznak J, Marzluf G, Schmidt T, Snyder L (ed), Methods for General and Molecular Microbiology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817497.ch7
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Image of FIGURE 2
FIGURE 2

Isolation of envelopes and related fractions from gram-negative bacteria.

Citation: Koval S, Sprott G. 2007. Cell Fractionation, p 108-137. In Reddy C, Beveridge T, Breznak J, Marzluf G, Schmidt T, Snyder L (ed), Methods for General and Molecular Microbiology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817497.ch7
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References

/content/book/10.1128/9781555817497.chap7
1. Altman, E.,, J.-R. Brisson,, and M. B. Perry. 1986. Structural studies of the capsular polysaccharide from Haemophilus pleuropneumoniae serotype 1. Biochem. Cell Biol. 64:707716.
2. Altman, E.,, M. B. Perry,, and J.-R. Brisson. 1989. Structure of the lipopolysaccharide antigenic O-chain produced by Actinobacillus pleuropneumoniae serotype 4 (ATCC 33378). Carbohydr. Res. 191:295303.
3. Ames, G. F.-L.,, C. Prody,, and S. Kustu. 1984. Simple, rapid, and quantitative release of periplasmic proteins by chloroform. J. Bacteriol. 160:11811183.
4. Archer, D. B.,, and N. R. King. 1984. Isolation of gas vesicles from Methanosarcina barkeri. J. Gen. Microbiol. 130:167172.
5. Austin, E. A.,, J.F. Graves,, L. A. Hite,, C. T. Parker,, and C. A. Schnaitman. 1990. Genetic analysis of lipopolysaccharide core biosynthesis by Escherichia coli K-12. J. Bacteriol. 172:53125325.
6. Austin, J. W.,, and R. G. E. Murray. 1987. The perforate component of the regularly-structured (RS) layer of Lampropedia hyalina. Can. J. Microbiol. 33:10391045.
7. Bakau, B.,, J. M. Brass,, and W. Boos. 1985. Ca2+-induced permeabilization of the Escherichia coli outer membrane: comparison of transformation and reconstitution of bindingprotein- dependent transport. J. Bacteriol. 163:6168.
8. Barber, E. J. 1976. Viscosity and density tables, p. 415418. In G. D. Fasman (ed.), Handbook of Biochemistry and Molecular Biology, 3rd ed., vol. 1. Physical and Chemical Data. CRC Press, Inc., Cleveland, OH.
9. Barrett, A. J. 1980. Introduction: the classification of proteinases. Ciba Found. Symp. 75:113.
10. Beaman, T. C.,, and P. Gerhardt. 1986. Heat resistance of bacterial spores correlated with protoplast dehydration, mineralization, and thermal adaptation. Appl. Environ. Microbiol. 52:12421246.
11. Beaman, T. C.,, J. T. Greenamyre,, T. R. Corner,, H. S. Pankratz,, and P. Gerhardt. 1982. Bacterial spore heat resistance correlated with water content, wet density, and protoplast/sporoplast volume ratio. J. Bacteriol. 150:870877.
12. Beveridge, T. J. 1979. Surface arrays on the wall of Sporosarcina urea. J. Bacteriol. 139:10391048.
13. Beveridge, T. J.,, G. B. Patel,, B. J. Harris,, and G. D. Sprott. 1986. The ultrastructure of Methanothrix concilii, a mesophilic aceticlastic methanogen. Can. J. Microbiol. 32:703710.
14. Beveridge, T. J.,, M. Stewart,, R. J. Doyle,, and G. D. Sprott. 1985. Unusual stability of the Methanospirillum hungatei sheath. J. Bacteriol. 162:728737.
15. Beynon, L. M.,, M. Moreau,, J. C. Richards,, and M. B. Perry. 1991. Structure of the O-antigen of Actinobacillus pleuropneumoniae serotype 7 lipopolysaccharide. Carbohydr. Res. 209:225238.
16. Bingle, W. H.,, J. L. Doran,, and W. J. Page. 1984. Regular surface layer of Azotobacter vinelandii. J. Bacteriol. 159:251259.
17. Blakemore, R. P. 1982. Magnetotactic bacteria. Annu. Rev. Microbiol. 36:217238.
18. Bohrmann, B.,, W. Villiger,, R. Johansen,, and E. Kellenberger. 1991. Coralline shape of the bacterial nucleoid after cryofixation. J. Bacteriol. 173:31493158.
19. Brinton, C. C.,, A. Buzzell,, and M. A. Lauffer. 1954. Electrophoresis and phage susceptibility on a filamentproducing variant of the E. coli B bacterium. Biochim. Biophys. Acta 15:533542.
20. Bush, J. W. 1985. Enzymatic lysis of the pseudomureincontaining methanogen Methanobacterium formicicum. J. Bacteriol. 163:2736.
21. Chen, W.,, K. Ohmiya,, and S. Shimizu. 1986. Protoplast formation and regeneration of dehydrodivanillin-degrading strains of Fusobacterium varium and Enterococcus faecium. Appl. Environ. Microbiol. 52:612616.
22. Clark, J. E.,, H. Beegen,, and H. G. Wood. 1985. Isolation of polyphosphate from Propionibacterium shermanii. Fed. Proc. 44:1079.
23. Coulton, J. W.,, and R. G. E. Murray. 1977. Membraneassociated components of the bacterial flagellar apparatus. Biochim. Biophys. Acta 465:290310.
24. Cunha, S.,, T. Odijk,, E. Suleymanoglu,, and C. L. Woldringh. 2001. Isolation of the Escherichia coli nucleoid. Biochimie 83:149154.
25. Cunningham, T. M.,, E. M. Walker,, J. N. Miller,, and M. A. Lovett. 1988. Selective release of the Treponema pallidum outer membrane and associated polypeptides with Triton X-114. J. Bacteriol. 170:57895796.
26. Darveau, R. P.,, and R. E. W. Hancock. 1983. Procedure for isolation of bacterial lipopolysaccharides from both smooth and rough Pseudomonas aeruginosa and Salmonella typhimurium strains. J. Bacteriol. 155:831838.
27. Davis, R. P.,, and J. E. Harris. 1985. Spontaneous protoplast formation by Methanosarcina barkeri. J. Gen. Microbiol. 131:14811486.
28. De Pamphilis, M. L.,, and J. Adler. 1971. Purification of intact flagella from Escherichia coli and Bacillus subtilis. J. Bacteriol. 105:376383.
29. Dubreuil, J. D.,, S. M. Logan,, S. Cubbage,, D. NiEidhin,, W. D. McCubbin,, C. M. Kay,, T. J. Beveridge,, F. G. Ferris,, and T. J. Trust. 1988. Structural and biochemical analysis of a surface array protein of Campylobacter fetus. J. Bacteriol. 170:41654173.
30. Dunn, R. M.,, M. J. Munster,, R. J. Sharp,, and B. N. Dancer. 1987. A novel method for regenerating the protoplasts of thermophilic bacilli. Arch. Microbiol. 146:323326.
31. Dupont, C.,, and A. J. Clarke. 1991. Dependence of lysozyme-catalysed solubilization of Proteus mirabilis peptidoglycan on the extent of O-acetylation. Eur. J. Biochem. 195:763769.
32. Easterbrook, K. B., 1989. Spinate bacteria, p. 19911993. In J.G. Holt (ed.), Bergey’s Manual of Systematic Bacteriology, vol. 3. The Williams and Wilkins Co., Baltimore, MD.
33. Easterbrook, K. B.,, and R. W. Coombs. 1976. Spinin: the subunit protein of bacterial spinae. Can. J. Microbiol. 22:438440.
34. Easterbrook, K. B.,, and S. Sperker. 1982. Physiological controls of bacterial spinae production in complex medium and their value as indicators of spina function. Can. J. Microbiol. 28:130136.
35. Elhardt, D.,, and A. Böck. 1982. An in vitro polypeptide synthesizing system from methanogenic bacteria: sensitivity to antibiotics. Mol. Gen. Genet. 188:128134.
36. Filip, C.,, G. Fletcher,, J. L. Wulff,, and C. F. Earhart. 1977. Solubilization of the cytoplasmic membrane of Escherichia coli by the ionic detergent sodium lauryl sarcosinate. J. Bacteriol. 115:717722.
37. Ford, T. C.,, and J. M. Graham. 1991. An Introduction to Centrifugation. BIOS Scientific Publishers Ltd., Oxford, United Kingdom.
38. Galanos, C.,, and O. Luderitz. 1975. Electrodialysis of lipopolysaccharides and their conversion to uniform salt form. Eur. J. Biochem. 54:603610.
39. Galanos, C.,, O. Lüderitz,, and O. Westphal. 1969. A new method for the extraction of R lipopolysaccharides. Eur. J. Biochem. 9:245249.
40. Gilleland, H. E., Jr.,, J. D. Stinnett,, I. L. Roth,, and R. G. Eagon. 1973. Freeze-etch study of Pseudomonas aeruginosa: localization within the cell wall of an ethylenediaminetetraacetate- extractable component. J. Bacteriol. 113:417432.
41. Gorby, Y. A.,, T. J. Beveridge,, and R. P. Blakemore. 1988. Characterization of the bacterial magnetosome membrane. J. Bacteriol. 170:834841.
42. Gould, G. W. 1971. Methods for studying bacterial spores. Methods Microbiol. 6A:361381.
43. Gray, G. W.,, and S. G. Wilkinson. 1965. The effect of ethylenediaminetetraacetic acid on the cell walls of some gram-negative bacteria. J. Gen. Microbiol. 39:385399.
44. Gruter, L.,, and R. Laufs. 1991. Protoplast transformation of Staphylococcus epidermidis. J. Microbiol. Methods 13:299304.
45. Hamilton, M. G. 1971. Isodensity equilibrium centrifugation of ribosomal particles; the calculation of the protein content of ribosomes and other ribonucleoproteins from buoyant density measurements. Methods Enzymol. 20:512521.
46. Hancock, I. C.,, and I. R. Poxton,. 1988. Isolation and purification of cell walls, p. 5565. In I. C. Hancock, and I. R. Poxton (ed.), Bacterial Cell Surface Techniques. John Wiley & Sons Ltd., Toronto, Canada.
47. Hancock, I. C.,, and I. R. Poxton,. 1988. Teichoic acids and other accessory carbohydrates from Gram-positive bacteria, p. 7988. In I. C. Hancock, and I. R. Poxton (ed.), Bacterial Cell Surface Techniques. John Wiley & Sons Ltd., Toronto, Canada.
48. Hancock, I. C.,, and I. R. Poxton,. 1988. Isolation of exopolysaccharides, p. 121125. In I. C. Hancock, and I. R. Poxton (ed.), Bacterial Cell Surface Techniques. John Wiley & Sons Ltd., Toronto, Canada.
49. Hancock, R. E. W.,, and H. Nikaido. 1978. Outer membranes of Gram-negative bacteria. XIX. Isolation from Pseudomonas aeruginosa PAO1 and use in reconstitution and definition of the permeability barrier. J. Bacteriol. 136:381390.
50. Harold, F. M. 1963. Accumulation of inorganic polyphosphate in Aerobacter aerogenes. I. Relationship to growth and nucleic acid synthesis. J. Bacteriol. 86:216221.
51. Harwood, C. R.,, and S. M. Cutting. 1990. Molecular Biological Methods for Bacillus. John Wiley & Sons, Inc., New York, NY.
52. Hash, J. H.,, and M. V. Rothlauf. 1967. The N,Odiacetylmuramidase of Chalaropsis species. J. Biol. Chem. 242:55865590.
53. Heckels, J. E.,, and M. Virji,. 1988. Detection and preparation of surface appendages, p. 6772. In I. C. Hancock, and I. R. Poxton (ed.), Bacterial Cell Surface Techniques. John Wiley & Sons Ltd., Toronto, Canada.
54. Helenius, A.,, D. R. McCaslin,, E. Fries,, and C. Tanford. 1979. Properties of detergents. Methods Enzymol. 56:734749.
55.. Heppel, L. A., 1971. The concept of periplasmic enzymes, p. 223247. In L. I. Rothfield (ed.), Structure and Function of Biological Membranes. Academic Press, Inc., New York, NY.
56. Hitchcock, P. J.,, and T. M. Brown. 1983. Morphological heterogeneity among Salmonella lipopolysaccharide chemotypes in silver-stained polyacrylamide gels. J. Bacteriol. 154:269277.
57. Hobot, J. A.,, E. Carlemalm,, W. Villiger,, and E. Kellenberger. 1984. Periplasmic gel: new concept resulting from the reinvestigation of bacterial cell envelope ultrastructure by new methods. J. Bacteriol. 160:143152.
58. Holloway, P. W. 1973. A simple procedure for the removal of Triton X-100 from protein samples. Anal. Biochem. 53:304308.
59. Hughes, D. E.,, J. W. T. Wimpenny,, and D. Lloyd,. 1971. The disintegration of microorganisms, p. 154. In J. R. Norris, and D. W. Ribbons (ed.), Methods in Microbiology, vol. 5B. Academic Press, Inc., New York, NY.
60. Jann, B.,, and K. Jann. 1990. Structure and biosynthesis of the capsular antigens of Escherichia coli, p. 19-42. Curr. Top. Microbiol. Immunol. 150:1942.
61. Jarrell, K. F.,, J. R. Colvin,, and G. D. Sprott. 1982. Spontaneous protoplast formation in Methanobacterium bryantii. J. Bacteriol. 149:346353.
62. Jarrell, K. F.,, D. Faguy,, A. M. Hebert,, and M. L. Kalmokoff. 1992. A general method of isolating high molecular weight DNA from methanogenic archaea (archaebacteria). Can. J. Microbiol. 38:6568.
63. Jarrell, K. F.,, and G. D. Sprott. 1984. Formation and regeneration of Halobacterium protoplasts. Curr. Microbiol. 10:147152.
64. Jennings, H. J. 1990. Capsular polysaccharides as vaccine candidates. Curr. Top. Microbiol. Immunol. 150:97127.
65. Johannsen, L.,, H. Labischinski,, B. Reinicke,, and P. Giesbrecht. 1983. Changes in the chemical structure of walls of Staphylococcus aureus grown in the presence of chloramphenicol. FEMS Microbiol. Lett. 16:313316.
66. Jollès, P. 1969. Lysozymes: a chapter of molecular biology. Angew. Chem. Int. Ed. 8:227294.
67. Jussofie, A.,, F. Mayer,, and G. Gottschalk. 1986. Methane formation from methanol and molecular hydrogen by protoplasts of new methanogen isolates and inhibition by dicyclohexylcarbodiimide. Arch. Microbiol. 146:245249.
68. Kaback, H. R. 1971. Bacterial membranes. Methods Enzymol. 22:99120.
69. Kalmokoff, M. L.,, K. F. Jarrell,, and S. F. Koval. 1988. Isolation of flagella from the archaebacterium Methanococcus voltae by phase separation with Triton X-114. J. Bacteriol. 170:17521758.
70. Kamekura, M.,, D. Oesterhelt,, R. Wallace,, P. Anderson,, and D. J. Kushner. 1988. Lysis of halobacteria in Bacto- Peptone by bile acids. Appl. Environ. Microbiol. 54:990995.
71. Kandler, O.,, and H. König. 1985. Cell envelopes of archaebacteria. Bacteria 8:413457.
72. Karch, H.,, H. Leying,, K.-H. Buscher,, H.-P. Kroll,, and W. Opferkuch. 1985. Isolation and separation of physiochemically distinct fimbrial types expressed on a single culture of Escherichia coli O7:K1:H6. Infect. Immun. 47:549554.
73. Kates, M.,, S. C. Kushwaha,, and G. D. Sprott. 1982. Lipids of purple membrane from extreme halophiles and of methanogenic bacteria. Methods Enzymol. 88:98111.
74. Kiener, A.,, H. König,, J. Winter,, and T. Leisinger. 1987. Purification and use of Methanobacterium wolfei pseudomurein endopeptidase for lysis of Methanobacterium thermoautotrophicum. J. Bacteriol. 169:10101016.
75. Kobayashi, H.,, J. van Brunt,, and F. M. Harold. 1978. ATP-linked calcium transport in cells and membrane vesicles of Streptococcus faecalis. J. Biol. Chem. 253:20852092.
76. Konig, H.,, and K. O. Stetter. 1986. Studies on archaebacterial S-layers. Syst. Appl. Microbiol. 7:300309.
77. Korch, C.,, S. Ovreb,, and K. Kleppe. 1976. Envelopeassociated folded chromosomes from Escherichia coli: variations under different physiological conditions. J. Bacteriol. 127:904916.
78. Kotzian, S.,, V. Kreis-Kleinschmidt,, T. Krafft,, O. Klimmek,, J. M. Macy,, and A. Kröger. 1996. Properties of a Wolinella succinogenes mutant lacking periplasmic sulfide dehydrogenase (Sud). Arch. Microbiol. 165:6568.
79. Koval, S. F. 1988. Paracrystalline protein surface arrays on bacteria. Can. J. Microbiol. 34:407414.
80. Koval, S. F.,, and K. F. Jarrell. 1987. Ultrastructure and biochemistry of the cell wall of Methanococcus voltae. J. Bacteriol. 169:12981306.
81. Koval, S. F.,, and R. G. E. Murray. 1984. The isolation of surface array proteins from bacteria. Can. J. Biochem. Cell Biol. 62:11811189.
82. Kulaev, I. S. 1975. Biochemistry of inorganic polyphosphates. Rev. Physiol. Biochem. Pharmacol. 73:131157.
83. Kurland, C. G. 1971. Purification of ribosomes from Escherichia coli. Methods Enzymol. 20:379381.
84. Kushwaha, S. C.,, M. Kates,, and W. G. Martin. 1975. Characterization and composition of the purple and red membrane from Halobacterium cutirubrum. Can. J. Microbiol. 53:284292.
85. Kushwaha, S. C.,, M. Kates,, and W. Stoeckenius. 1976. Comparison of the purple membrane from Halobacterium cutirubrum and Halobacterium halobium. Biochim. Biophys. Acta 426:703710.
86. Laddaga, R. A.,, and R. A. MacLeod. 1982. Effects of wash treatments on the ultrastructure and lysozyme penetrability of the outer membrane of various marine and two terrestrial gram-negative bacteria. Can. J. Microbiol. 28:318324.
87. Langworthy, T. A., 1978. Membranes and lipids of extremely thermoacidophilic microorganisms, p. 1130. In S. M. Friedman (ed.), Biochemistry of Thermophily. Academic Press, Inc., New York, NY.
88. Leive, L. 1965. Release of lipopolysaccharide by EDTA treatment of Escherichia coli. Biochem. Biophys. Res. Commun. 21:290296.
89. Lesse, A. J.,, A. A. Campagnari,, W. E. Bittner,, and M. A. Apicella. 1990. Increased resolution of lipopolysaccharides and lipooligosaccharides utilizing tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis. J. Immunol. Methods 126:109117.
90. Li, N.,, and M. C. Cannon. 1998. Gas vesicle genes identified in Bacillus megaterium and functional expression in Escherichia coli. J. Bacteriol. 180:24502458.
91. Lindsay, J. A.,, T. C. Beaman,, and P. Gerhardt. 1985. Protoplast water content of bacterial spores determined by buoyant density sedimentation. J. Bacteriol. 163:735737.
92. Lu, W.,, A. P. Wood,, and D. P. Kelly. 1983. An enzymatic lysis procedure for the assay of enzymes in Thiobacillus A2. Microbios 38:171176.
93. Lugtenberg, B.,, and L. van Alphen. 1983. Molecular architecture and functioning of the outer membrane of Escherichia coli and other Gram-negative bacteria. Biochim. Biophys. Acta 737:51115.
94. MacDonald, R. E.,, and J. K. Lanyi. 1975. Light-induced leucine transport in Halobacterium halobium envelope vesicles: a chemiosmotic study. Biochemistry 14:28822889.
95. MacLeod, R. A. 1985. Marine microbiology far from the sea. Annu. Rev. Microbiol. 39:120.
96. Manoharan, R.,, E. Ghiamati,, R. A. Dalterio,, K. A. Britton,, W. H. Nelson,, and J. F. Sperry. 1990. UV resonance Raman spectra of bacteria, bacterial spores, protoplasts and calcium dipicolinate. J. Microbiol. Methods 11:115.
97. Marvin, H. J. P.,, and B. Witholt. 1987. A highly efficient procedure for the quantitative formation of intact and viable-lysozyme spheroplasts from Escherichia coli. Anal. Biochem. 164:320330.
98. Matheson, A. T.,, M. Yaguchi,, W. E. Balch,, and R. S. Wolfe. 1980. Sequence homologies in the N-terminal region of the ribosomal “A” proteins from Methanobacterium thermoautotrophicum and Halobacterium cutirubrum. Biochim. Biophys. Acta 626:162169.
99. Mayer, F.,, A. Jussofie,, M. Salzmann,, M. Lübben,, M. Rohde,, and G. Gottschalk. 1987. Immunoelectron microscopic demonstration of ATPase on the cytoplasmic membrane of the methanogenic bacterium strain GÖ1. J. Bacteriol. 169:23072309.
100. Meakin, S. A.,, J. H. E. Nash,, W. D. Murray,, K. J. Kennedy,, and G. D. Sprott. 1991. A generally applicable technique for the extraction of restrictable DNA from methanogenic bacteria. J. Microbiol. Methods 14:119126.
101. Messner, P.,, and U. B. Sleytr,. 1988. Separation and purification of S-layers from Gram-positive and Gramnegative bacteria, p. 97104. In I. Hancock, and I. Poxton (ed.), Bacterial Cell Surface Techniques. John Wiley & Sons Ltd., Toronto, Canada.
102. Miura, T.,, and S. Mizushima. 1968. Separation by density gradient centrifugation of two types of membranes from spheroplast membranes of Escherichia coli K12. Biochim. Biophys. Acta 150:159161.
103. Moore, P. B. 1979. The preparation of deuterated ribosomal materials for neutron scattering. Methods Enzymol. 59:639655.
104. Morii, H.,, and Y. Koga. 1992. An improved assay for a pseudomurein-degrading enzyme from Methanobacterium wolfei and the protoplast formation of Methanobacterium thermoautotrophicum by the enzyme. J. Ferment. Bioeng. 73:610.
105. Nilles, M. L.,, A. W. Williams,, E. Skrzypek,, and S. C. Straley. 1997. Yersinia pestis LcrV forms a stable complex with LcrG and may have a secretion-related regulatory role in the low-Ca2+ response. J. Bacteriol. 179:13071316.
106. Nossal, N. G.,, and L. A. Heppel. 1966. The release of enzymes by osmotic shock from Escherichia coli in exponential phase. J. Biol. Chem. 241:30553062.
107. Offner, S.,, A. Hofacker,, G. Wanner,, and F. Pfeifer. 2000. Eight of fourteen gvp genes are sufficient for formation of gas vesicles in halophilic archaea. J. Bacteriol. 182:43284336.
108. Offner, S.,, G. Wanner,, and F. Pfeifer. 1996. Functional studies of the gvpACNO operon of Halobacterium salinarium reveal that the GvpC protein shapes gas vesicles. J. Bacteriol. 178:20712078.
109. Osborn, M. J.,, and R. Munson. 1974. Separation of the inner (cytoplasmic) and outer membranes of gram negative bacteria. Methods Enzymol . 31A:642653.
110. Paranchych, W.,, P. A. Sastry,, L. S. Frost,, M. Carpenter,, G. D. Armstrong,, and T. H. Watts. 1979. Biochemical studies on pili isolated from Pseudomonas aeruginosa strain PAO. Can. J. Microbiol. 25:11751181.
111. Penn, C. W.,, A. Cockayne,, and M. J. Bailey. 1985. The outer membrane of Treponema pallidum: biological significance and biochemical properties. J. Gen. Microbiol. 131:23492357.
112. Pertoft, H.,, T. C. Laurent,, R. Seljelid,, G. Akerstrom,, L. Kagedal,, and M. Hirtenstein,. 1979. The use of density gradients of PercollR for the separation of biological particles, p. 6772. In H. Peeters (ed.), Separation of Cells and Subcellular Elements. Pergamon Press, Toronto, Canada.
113. Peterson, G. L. 1977. A simplification of the protein assay method of Lowry et al. which is more generally applicable. Anal. Biochem. 83:346356.
114. Petrov, V. V.,, V. Y. Artzatbanov,, E. N. Ratner,, A. I. Severin,, and I. S. Kulaev. 1991. Isolation, structural and functional characterization of Staphylococcus aureus protoplasts obtained using lysoamidase. Arch. Microbiol. 155:549553.
115. Price, C.A. 1982. Centrifugation in Density Gradients. Academic Press, New York, NY.
116. Rheinberger, H.-J.,, U. Geigenmüller,, M. Wedde,, and K. H. Nierhaus. 1988. Parameters for the preparation of Escherichia coli ribosomes and ribosomal subunits active in tRNA binding. Methods Enzymol. 164:658670.
117. Rickwood, D. (ed.). 1984. Centrifugation: a Practical Approach, 2nd ed. IRL Press, Oxford, United Kingdom.
118. Ross, J. W. 1963. Continuous-flow mechanical cell disintegrator. Appl. Microbiol. 11:3335.
119. Salton, M. R. J. 1957. The properties of lysozyme and its action on microorganisms. Bacteriol. Rev. 21:8299.
120. Salton, M. R. J. 1974. Isolation of cell walls from Grampositive bacteria. Methods Enzymol. 31:653667.
121. Sambrook, J.,, E. F. Fritsch,, and T. Maniatis. 1989. Molecular Cloning: a Laboratory Manual, 2nd ed. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY.
122. Scherer, P. A.,, and H.-P. Bochem. 1983. Ultrastructural investigation of 12 Methanosarcinae and related species grown on methanol for occurrence of polyphosphate-like inclusion. Can. J. Microbiol. 29:11901199.
123. Schmidt, K. 1980. A comparative study on the composition of chlorosomes (chlorobium vesicles) and cytoplasmic membranes from Chloroflex aurantiacus strain OK-70- f1 and Chlorobium thiosulfatophilum strain 6230. Arch. Microbiol. 124:2131.
124. Schnaitman, C. A. 1970. Protein composition of the cell wall and cytoplasmic membrane of Escherichia coli. J. Bacteriol. 104:890901.
125. Schnaitman, C. A. 1971. Solubilization of the cytoplasmic membrane of Escherichia coli by Triton X-100. J. Bacteriol. 108:545552.
126. Schüler, D.,, and R. B. Frankel. 1999. Bacterial magnetosomes: microbiology, biomineralization and biotechnological applications. Appl. Microbiol. Biotechnol. 52:464473.
127. Schulman, H.,, and E. P. Kennedy. 1979. Localization of membrane-derived oligosaccharides in the outer envelope of Escherichia coli and their occurrence in other gram-negative bacteria. J. Bacteriol. 137:686688.
128. Sharrock, W. J.,, and J. C. Rabinowitz. 1979. Fractionation of ribosomal particles from Bacillus subtilis. Methods Enzymol. 59:371382.
129. Siegel, J. L.,, S. F. Hurst,, E. S. Liberman,, S. E. Coleman,, and A. S. Bleiweis. 1981. Mutanolysin-induced spheroplasts of Streptococcus mutans are true protoplasts. Infect. Immun. 31:808815.
130. Simon, R. D. 1981. Morphology and protein composition of gas vesicles from wild type and gas vacuole defective strains of Halobacterium salinarium strain 5. J. Gen. Microbiol. 125:103111.
131. Sleytr, W. B.,, and P. Messner. 1988. Crystalline surface layers in procaryotes. J. Bacteriol. 170:28912897.
132. Spizizen, J. 1962. Preparation and use of protoplasts. Methods Enzymol. 5:122134.
133. Sprott, G. D.,, J. R. Colvin,, and R. C. McKellar. 1979. Spheroplasts of Methanospirillum hungatei formed upon treatment with dithiothreitol. Can. J. Microbiol. 25:730738.
134. Sprott, G. D.,, and K. F. Jarrell. 1981. K+, Na+, and Mg+ content and permeability of Methanospirillum hungatei and Methanobacterium thermoautotrophicum. Can. J. Microbiol. 27:444451.
135. Sprott, G. D.,, K. M. Shaw,, and K. F. Jarrell. 1983. Isolation and chemical composition of the cytoplasmic membrane of the archaebacterium Methanospirillum hungatei. J. Biol. Chem. 258:40264031.
136. Staechelin, T.,, and D. R. Maglott. 1971. Preparation of Escherichia coli ribosomal subunits active in polypeptide synthesis. Methods Enzymol. 20:449456.
137. Steinberg, W. 1974. Properties and developmental roles of the lysyl- and tryptophanyl-transfer ribonucleic acid synthetase of Bacillus subtilis: common genetic origin of the corresponding spore and vegetative enzymes. J. Bacteriol. 118:7082.
138. Stoeckenius, W.,, and R. Rowen. 1967. A morphological study of Halobacterium halobium and its lysis in media of low salt concentration. J. Cell Biol. 34:365393.
139. Stonington, O. G.,, and D. E. Pettijohn. 1971. The folded genome of Escherichia coli isolated in a protein-DNARNA complex. Proc. Natl. Acad. Sci. USA 68:69.
140. Sykes, J., 1971. Centrifugation techniques for the isolation and characterization of sub-cellular components from bacteria, p. 55207. In J. R. Norris, and D. W. Ribbons (ed.), Methods in Microbiology, vol. 5B. Academic Press, Inc., New York, NY.
141. Thom, J. R.,, and L. L. Randall. 1988. Role of the leader peptide of maltose-binding protein in two steps of the export process. J. Bacteriol. 170:56545661.
142. Thomas, N. A.,, S. L. Bardy,, and K. F. Jarrell. 2001. The archaeal flagellum: a different kind of prokaryotic motility structure. FEMS Microbiol. Lett. 25:147174.
143. Thompson, B. G.,, R. G. E. Murray,, and J. F. Boyce. 1982. The association of the surface array and the outer membrane of Deinococcus radiodurans. Can. J. Microbiol. 28:10811088.
144. Tisa, L. S.,, T. Koshikawa,, and P. Gerhardt. 1982. Wet and dry bacterial spore densities determined by buoyant sedimentation. Appl. Environ. Microbiol. 43:13071310.
145. Vainshtein, M. B.,, N. E. Suzina,, E. B. Kudryashova,, E. V. Ariskina,, and V. V. Sorokin. 1998. On the diversity of magnetotactic bacteria. Microbiology 67:670676.
146. Vandenbergh, P. A.,, R. E. Bawdon,, and R. S. Berk. 1979. Rapid test for determining the intracellular rhodanese activity of various bacteria. Int. J. Syst. Bacteriol. 29:339344.
147. Virgilio, R.,, C. Gonzalez,, N. Muñoz,, T. Cabezon,, and S. Mendoza. 1970. Staphylococcus aureus protoplasting induced by D-cycloserine. J. Bacteriol. 104:13861387.
148. Walker, J. E.,, P. K. Hayes,, and A. E. Walsby. 1984. Homology of gas vesicle proteins in Cyanobacteria and Halobacteria. J. Gen. Microbiol. 130:27092715.
149. Walsby, A. E. 1974. The isolation of gas vesicles from blue-green algae. Methods Enzymol. 31:678686.
150. Ward, J. B. 1981. Teichoic and teichuronic acids: biosynthesis, assembly, and location. Microbiol. Rev. 45:211243.
151. Warth, A. D. 1978. Relationship between the heat resistance of spores and the optimum and maximum growth temperature of Bacillus species. J. Bacteriol. 134:699705.
152. Weil, C. F.,, D. S. Cram,, B. A. Sherf,, and J. N. Reeve. 1988. Structure and comparative analysis of the genes encoding component C of methyl coenzyme M reductase in the extremely thermophilic archaebacterium Methanothermus fervidus. J. Bacteriol. 170:47184726.
153. Westphal, O.,, and K. Jann. 1965. Bacterial lipopolysaccharides. Methods Carbohydr. Chem. 5:8391.
154. Wilkinson, S. G.,, L. Galbraith,, and G. A. Lightfoot. 1973. Cell walls, lipids, and lipopolysaccharides of Pseudomonas species. Eur. J. Biochem. 33:158174.
155. Worcel, A.,, and E. Burgi. 1972. On the structure of the folded chromosome of Escherichia coli. J. Mol. Biol. 71:127147.
156. Work, E., 1971. Cell walls, p. 361418. In J. R. Norris, and D. W. Ribbons (ed.), Methods in Microbiology, vol. 5A. Academic Press, Inc., New York, NY.
157. Yabu, K.,, and S. Takahashi. 1977. Protoplast formation of selected Mycobacterium smegmatis mutants by lysozyme in combination with methionine. J. Bacteriol. 129: 16281631.
158. Yamada, H.,, N. Tsukagoshi,, and S. Udaka. 1981. Morphological alterations of cell wall concomitant with protein release in a protein-producing bacterium, Bacillus brevis 47. J. Bacteriol. 148:322332.
159. Yokomaku, D.,, N. Yamaguchi,, and M. Nasu. 2000. Improved direct viable count procedure for quantitative estimation of bacterial viability in freshwater environments. Appl. Environ. Microbiol. 66:55445548.
160. Zulauf, M.,, U. Fürstenberger,, M. Grabo,, P. Jäggi,, M. Regenass,, and J. P. Rosenbusch. 1989. Critical micellar concentrations of detergents. Methods Enzymol. 172:528538.

Tables

Generic image for table
TABLE 1

Some commonly used methods to lyse bacteria and archaea to form osmotically sensitive cells in hypotonic solutions

Lysozyme susceptibility follows growth in medium containing penicillin G.

Citation: Koval S, Sprott G. 2007. Cell Fractionation, p 108-137. In Reddy C, Beveridge T, Breznak J, Marzluf G, Schmidt T, Snyder L (ed), Methods for General and Molecular Microbiology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817497.ch7
Generic image for table
TABLE 2

Buoyant densities of fractions isolated from prokaryotic cells

Citation: Koval S, Sprott G. 2007. Cell Fractionation, p 108-137. In Reddy C, Beveridge T, Breznak J, Marzluf G, Schmidt T, Snyder L (ed), Methods for General and Molecular Microbiology, Third Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817497.ch7

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