The periplasm is a space bordered by two selective permeable barriers, i.e., biological membranes, which are the inner membrane (i.e., cytoplasmic membrane) and the outer membrane in Gram-negative bacteria. Strictly speaking, there is no periplasmic space in Gram-positive bacteria because there is only one biological membrane, the cytoplasmic membrane. A region termed "inner wall zone" (IWZ) has been observed between the cytoplasmic membrane and the mature cell wall.
The periplasm may constitute up to 40% of the total cell volume in Gram-negative species, whereas the IWZ is drastically smaller in Gram-positive species. Composition of the IWZ has not been as yet clearly determined.
Although the bacteria are conventionally divided into two main groups — Gram-positive and -negative, based upon their Gram-stain retention property — this classification system is ambiguous as it can refers to three distinct aspects (staining result, cell-envelope organization, taxonomic group), which do not necessarily coalesce for some bacterial species. However, although Gram-staining response of bacteria is an empirical criterion, its basis lies in the marked differences in the ultrastructure and chemical composition of the two main kinds of prokaryotic cells that are found in nature. These two kinds of cells are distinguished from each other based upon the presence or absence of an outer lipid membrane, which is a more reliable and fundamental characteristic of the bacterial cells.
All Gram-positive bacteria are bounded by a single unit lipid membrane; they generally contain a thick layer (20-80 nm) of peptidoglycan responsible for retaining the Gram-stain. A number of other bacteria which are bounded by a single membrane but stain Gram-negative due to either lack of the peptidoglycan layer (viz., mycoplasmas) or their inability to retain the Gram-stain due to their cell wall composition, also show close relationship to the gram-positive bacteria. For the bacterial (prokaryotic) cells that are bounded by a single cell membrane the term "mononderm bacteria" or "monoderm prokaryotes" has been proposed. In contrast to Gram-positive bacteria, all archetypical Gram-negative bacteria are bounded by a cytoplasmic membrane as well as an outer cell membrane; they contain only a thin layer of peptidoglycan (2-3 nm) between these membranes. The presence of both inner and outer cell membranes forms and define the periplasmic space or periplasmic compartment. These bacterial cells with two membranes have been designated as diderm bacteria. The distinction between the monoderm and diderm prokaryotes is supported by conserved signature indels in a number of important proteins (viz. DnaK, GroEL).
In diderm bacteria, the periplasm contains a thin cell wall composed of peptidoglycan. In addition, it includes solutes such as ions and proteins, which are involved in wide variety of functions ranging from nutrient binding, transport, folding, degradation, substrate hydrolysis, to peptidoglycan synthesis, electron transport, and alteration of substances toxic to the cell (xenobiotic metabolism). Importantly, the periplasm is devoid of ATP.
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