My NCBISign in to NCBISign Out US National Library of Medicine
National Institutes of Health 8550511
The permeability of the wall fabric of Escherichia coli and Bacillus subtilis.P Demchick and A L Koch Author information Copyright and License information Disclaimer Biological and Physical Sciences, Barton College, Wilson, North Carolina 27893, USA. Copyright notice This article has been cited by other articles in PMC.
To study the overall structure of the peptidoglycan fabric of the sacculi of gram-negative and gram-positive walls, actively growing cultures of Escherichia coli and Bacillus subtilis were treated with boiling sodium dodecyl sulfate solutions. The sacculi were then treated with enzymes to eliminate proteins and nucleic acids. These intact saccoli were probed with fluorescein-labeled dextrans with a range of known molecular weights. The penetration of the probes could be monitored by the negative-staining appearance in the fluorescence microscope. At several chosen times, the molecular weight fraction that allowed barely observable entry of the fluorescein-labeled probe and the molecular weight fraction that penetrated to achieve almost, but not quite, the concentration of probe in the solution external to the sacculi were determined. From three pairs of times and molecular weights that met one or the other of these two criteria, the effective pore size could be calculated. The minimum size of protein molecule that could diffuse through the pores was also calculated. Two mathematical models, which gave essentially the same results, were used to interpret the experimental data: one for the permeation of random coils through a surface containing holes and the other for rigid spheres diffusing through water-filled cylindrical pores. The mean estimate of the effective hole radius in walls from E. coli is 2.06 nm, and that of the effective hole size in walls from B. subtilis is 2.12 nm. These results are supported by experiments in which the loss of preloaded cells was monitored. Various fluorescein-labeled dextran samples were mixed with samples of intact cell walls, held for a long time, and then diluted. The efflux of the dextrans was monitored. Neither large nor small dextrans stained under these conditions. Only with dextran samples of a sufficiently small size were the sacculi filled during the preincubation period, and only with the largest of these could the probe not escape quickly. From the pore (or mesh) size, it can be concluded that the wall fabric of both organisms has few imperfections and that the major passageway is through the smallest possible pore, or "tessera," formed by the maximal cross-linking of the peptides from glycan chain to glycan chain compatible with the degree of rotational flexibility of the chains of repeating disaccharides of N-acetyl muramic acid and N-acetyl glucosamine. A tessera is composed of two chains of eight saccharides cross-linked by two octapeptides. The size of a globular hydrophilic molecule, if it did not bind to wall components, that could pass freely through the meshwork of an unstretched sacculus of either organism is roughly 25 kDa. We stress that this is only a rough estimate, and it may be possible for proteins of less than 50 kDa to pass through the native wall during normal growth conditions.
Full TextThe Full Text of this article is available as a PDF (232K).
These references are in PubMed. This may not be the complete list of references from this article.
- GERHARDT P, JUDGE JA. POROSITY OF ISOLATED CELL WALLS OF SACCHAROMYCES CEREVISIAE AND BACILLUS MEGATERIUM. J Bacteriol. 1964 Apr;87:945–951. [PMC free article] [PubMed]
- Granath KA, Kvist BE. Molecular weight distribution analysis by gel chromatography on Sephadex. J Chromatogr. 1967 May;28(1):69–81. [PubMed]
- Hughes RC, Thurman PF, Stokes E. Estimates of the porosity of Bacillus licheniformis and Bacillus subtilis cell walls. Z Immunitatsforsch Exp Klin Immunol. 1975 Jul;149(2-4):126–135. [PubMed]
- Kemper MA, Urrutia MM, Beveridge TJ, Koch AL, Doyle RJ. Proton motive force may regulate cell wall-associated enzymes of Bacillus subtilis. J Bacteriol. 1993 Sep;175(17):5690–5696. [PMC free article] [PubMed]
- Koch AL. The surface stress theory of microbial morphogenesis. Adv Microb Physiol. 1983;24:301–366. [PubMed]
- Koch AL. Shrinkage of growing Escherichia coli cells by osmotic challenge. J Bacteriol. 1984 Sep;159(3):919–924. [PMC free article] [PubMed]
- Koch AL, Doyle RJ. Inside-to-outside growth and turnover of the wall of gram-positive rods. J Theor Biol. 1985 Nov 7;117(1):137–157. [PubMed]
- Koch AL, Lane SL, Miller JA, Nickens DG. Contraction of filaments of Escherichia coli after disruption of cell membrane by detergent. J Bacteriol. 1987 May;169(5):1979–1984. [PMC free article] [PubMed]
- Koch AL, Woeste S. Elasticity of the sacculus of Escherichia coli. J Bacteriol. 1992 Jul;174(14):4811–4819. [PMC free article] [PubMed]
- Kurtzhals P, Larsen C, Johansen M. High-performance size-exclusion chromatographic procedure for the determination of fluoresceinyl isothiocyanate dextrans of various molecular masses in biological media. J Chromatogr. 1989 Jun 30;491(1):117–127. [PubMed]
- RENKIN EM. Filtration, diffusion, and molecular sieving through porous cellulose membranes. J Gen Physiol. 1954 Nov 20;38(2):225–243. [PMC free article] [PubMed]
- Scherrer R, Berlin E, Gerhardt Density, porosity, and structure of dried cell walls isolated from Bacillus megaterium and Saccharomyces cerevisiae. J Bacteriol. 1977 Feb;129(2):1162–1164. [PMC free article] [PubMed]
- Scherrer R, Gerhardt P. Molecular sieving by the Bacillus megaterium cell wall and protoplast. J Bacteriol. 1971 Sep;107(3):718–735. [PMC free article] [PubMed]
Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)