Biochemistry and Molecular Biology - Belyasova, N. A. 2002

Structure and Functions of Cellular Components
Cellular Polysaccharides
Structure of Prokaryotic Cell Walls

Most prokaryotes have a rigid, multilayered Cell wall On the surface of their Plasma Membrane, which serves to mechanically protect The Cell and maintain its shape. The cell wall may be separated from the membrane by a periplasmic space containing Enzymes and nutrient-binding Proteins. Unlike The Plasma Membrane, the cell wall is permeable to salts and low-molecular-weight compounds. The cell walls of most prokaryotes are primarily composed of the peptidoglycan murein. However, the Composition and Structure of cell walls vary significantly between the two major prokaryotic taxa (Gracilicutes and Firmicutes), which forms The basis of the differential Gram staining method for Bacteria.

Cell wall of Gram-positive bacteria. This is composed almost entirely of murein. This heteropolymer contains alternating residues of N-acetylglucosamine and N-acetylmuramic acid linked together by β(1→4)-glycosidic bonds. The murein molecule is unique in its structure because it contains D-Amino Acids, diaminopimelic acid, and a y-peptide bond (Fig. 5.4).

The unusual structure of murein is manifested primarily in the Features of the tetrapeptide moiety within N-acetylmuramic acid. The first position is occupied by L-Alanine, and the second by D-isoglutamic acid (or a minor amino acid). Isoglutamic acid forms a peptide bond with L-Lysine via its y-carboxyl group (y-peptide bond) rather than the usual a-carboxyl group. The third position in the tetrapeptide is occupied by L-lysine, which features a side-chain amino group, although it can be substituted with Other Amino Acids possessing a side-chain amino group (such as diaminopimelic or diaminobutyric acid) (Fig. 5.5). The final position in the tetrapeptide is always occupied by D-alanine. The presence of D-amino acids in murein confers enhanced resistance to Proteolytic Enzymes.

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Fig. 5.4. STRUCTURE OF THE monomeric unit of the murein molecule

Linear peptidoglycan chains, consisting of alternating disaccharide-peptide units (Fig. 5.4), form the backbone of murein. Its structure is further complicated by short Peptides that cross-link the polysaccharide chains. Numerous cross-links give rise to a single massive sacculus-like macromolecule known as the murein sacculus. These cross-links are most commonly pentaglycine peptides (bridges), although other amino acids may also be incorporated, such as L-alanine, L-Threonine, or L-Serine. During cross-link formation, the carboxyl group of one terminal Glycine residue in the pentaglycine bridge forms a peptide bond with the side-chain amino group of lysine within the tetrapeptide of N-acetylmuramic acid in one chain, while the free amino group of the other terminal residue in the pentaglycine bridge forms a peptide bond with the free carboxyl group of the terminal D-alanine residue in the N-acetylmuramic acid of the same or another murein chain (Fig. 5.6). Thus, a single tetrapeptide can participate in forming two cross-links between chains simultaneously via pentaglycine bridges.

The resulting complex murein network is reinforced by Teichoic Acids, which overlay or impregnate the peptidoglycan layers. Teichoic acids are chains of glycerol or the pentahydric alcohol ribitol molecules linked together by phosphodiester bridges (Fig. 5.5). A single teichoic acid molecule contains approximately 30 alcohol residues. Amino acid or sugar residues may also be incorporated into this chain. The hydroxyl groups of the alcohol constituents in teichoic acids serve to bind the peptidoglycan layers to one another, thereby imparting greater rigidity to the murein framework.

Fig. 5.5. Structure of individual components of Introduction/4.html">Prokaryotic Cell walls

Fig. 5.6. Structure of murein: NAG stands for the N-acetylglucosamine residue in the linear peptidoglycan chain; NAM stands for the N-acetylmuramic acid residue; (Gly)₅ represents the pentaglycine bridge linking amino acids within the tetrapeptide of N-acetylmuramic acid

Overall, the cell wall of Gram-positive bacteria contains about 40 layers of murein, which accounts for 30–70% of the dry weight of the cell wall. The wall maintains its structural integrity and cell shape even when the cellular contents are removed, for instance, by acid dissolution.

Cell wall of Gram-negative bacteria. The cell wall of Gram-negative bacteria has a more complex Organization than that of Gram-positive bacteria. Its main distinctive features are as follows: the murein network is single-layered and accounts for only 5–10% of the dry weight of the cell wall; teichoic acids are absent; and a lipid-rich outer membrane is present on The surface of the peptidoglycan layer (Fig. 5.7). Structurally, the murein of Gram-negative bacteria differs from that of Gram-positive bacteria by having a lower degree of cross-linking and a smaller variety of diamino acids.

The outer membrane constitutes a thick layer of the cell wall and is composed of lipopolysaccharides, Phospholipids, and proteins. The fundamental architecture of this layer closely resembles that of the plasma membrane. Immediately overlying the murein layer is a phospholipid layer, with molecules oriented such that their hydrophilic heads face the murein, while their hydrophobic tails face the lipopolysaccharide layer. Lipopolysaccharides serve as the primary Components of the outer membrane and possess an unusual structure. Their molecules consist of three regions: lipid A, the core oligosaccharide, and O-Antigens.

Lipid A, the backbone of this complex molecule, is composed of disaccharide units formed by two N-acetylglucosamine residues linked by β-(1→4)-glycosidic bonds, which repeat multiple times and are interconnected by phosphodiester bridges. Fatty acid residues are attached to the N-acetylglucosamine residues and, together with phospholipids, form a lipid bilayer. Protein molecules are embedded within The Lipid Bilayer formed by lipopolysaccharides and phospholipids. These proteins traverse the bilayer and function as transmembrane transport proteins responsible for the translocation of substances across the outer membrane. Externally, polysaccharide molecules are attached to lipid A; based on their length, these can be divided into two parts: the core (comprising 8–10 monosaccharide residues and exhibiting a conserved structure across almost all Gram-negative bacteria) and O-antigens. The latter represent the variable region of the outer Polysaccharides and most commonly consist of tetrasaccharide repeats that may occur up to 50 times within a single molecule (Fig. 5.7).

Fig. 5.7. Structure of the Gram-negative Bacterial cell wall

Unlike the acyl portion of lipid A, the core Oligosaccharides and O-antigen side chains are highly hydrophilic. Several sugars in the lipopolysaccharide are phosphorylated, giving the molecule an overall negative charge. To maintain the Stability of the lipopolysaccharide layer, Ca²⁺ ions are present in high concentrations. The O-side chains reside at the outermost surface of the cell, forming a so-called "molecular nap" or fuzzy layer. They serve as receptors for the adsorption of many Bacteriophages and act as the primary antigenic determinants of the Gram-negative cell wall. Furthermore, they are referred to as endotoxins because the virulence of bacterial Cells in animals is determined by the presence and type of O-side chains. Experiments involving the administration of isolated cell walls of specific bacterial strains to laboratory animals successfully replicate the symptoms of the corresponding disease, inducing Immunity in the animals against bacteria possessing those specific cell walls.

The outer membrane is anchored to the murein network by Lipoproteins, which attach to murein via diaminopimelic acid. Meanwhile, the lipophilic (hydrophobic) domains of the lipoproteins are embedded within the lipid bilayer, functioning as an anchor.

Various chemical agents can affect prokaryotic cell walls, notably widely used agents such as the enzyme Lysozyme and the antibiotic penicillin. The mechanisms of these actions will be discussed in subsequent sections. A common outcome of these and similar agents is that, in hypertonic environments, they lead to The formation of protoplasts (forms completely devoid of a cell wall) or spheroplasts (forms partially lacking a cell wall). These structures are osmotically fragile (undergoing lysis in hypotonic environments) and are capable of regenerating their cell wall under suitable conditions. Protoplasts and spheroplasts are widely utilized in Genetic Engineering for Cell Fusion and transformation experiments to generate hybrid bacteria.