Biochemistry and Molecular Biology - Belyasova N.A. 2002

Structure and Functions of Cellular Components
Biomembranes
Transport of Substances Across Membranes

Cellular activity is closely linked to the continuous exchange of contents between The Cell and its external environment. Similarly, substances constantly move within the cell between its Organelles and compartments. All of these processes involve overcoming a primary barrier to molecular transport—the membrane that encloses either the organelle or the cell itself. It is important to bear in mind that the principal function of Biomembranes is the selective transport of various substances and ions. The potential mechanisms of membrane transport can be divided into four main types: passive diffusion, Facilitated Diffusion, Active Transport, and cytosis.

Passive diffusion. This is a transport process in which substances move across membranes from a region of higher concentration to a region of lower concentration (along a chemical gradient) without the involvement of transport Proteins or the expenditure of energy. This pathway is utilized for the Transport of Small, uncharged molecules across the membrane, such as gas molecules, certain anesthetics, and Water. To cross the bilayer, a molecule must overcome surface tension at the membrane interface, penetrate the bilayer, diffuse across it, and emerge on the opposite side by once again surmounting the phase-boundary energy barrier. This accounts for the selective permeability of The Lipid Bilayer to small non-electrolyte molecules. Remarkably, water penetrates membranes with great ease and speed: it has been demonstrated that a water molecule takes a mere 1 µs to cross the bilayer. To account for this phenomenon, recent hypotheses—supported by certain experimental data—suggest that either some form of proteinaceous water-conducting channels do exist within membranes, or water molecules exploit local structural defects in the lipid bilayers.

The Movement of water molecules alone across a semipermeable membrane can be viewed as a specific type of diffusion known as osmosis. Osmosis is defined as the net movement of water molecules from a region of high water potential and low solute concentration to a region of low water potential and high solute concentration (Fig. 4.9). In this scenario, water molecules will flow from a hypotonic solution into a hypertonic solution until equilibrium is reached and both solutions become isotonic relative to each other.

To quantify the decrease in water potential caused by the presence of solutes, the term “osmotic pressure” is employed. Osmotic pressure refers to the pressure that must be applied to a solution to halt the osmotic influx of water through a semipermeable membrane. An increase in solute concentration raises the osmotic pressure and lowers The water potential of the solution.

The movement of water across cellular Plasma Membranes in accordance with the laws of osmosis poses significant challenges for living organisms, particularly aquatic species. Consequently, osmoregulation (maintaining a constant water potential within the cell) is a vital aspect of the physiological activity of most organisms, frequently consuming a substantial fraction of the energy stored by the cell.

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Fig. 4.9. Movement of water molecules in accordance with the laws of osmosis. Solution A has a higher glucose concentration than solution B and is therefore hypertonic to it. Conversely, solution B is hypotonic relative to solution A. The osmotic movement of water results in the equalization of both water and glucose concentrations, rendering the solutions isotonic to each other.

The rate of molecular diffusion across membranes depends on numerous factors: the solubility of the substance in the membrane, the membrane diffusion coefficient, and the concentration gradient between the extracellular and intracellular environments (Fig. 4.10).

Facilitated diffusion. This type of transport is mediated by transport proteins down an electrochemical gradient (accounting for both electrical potential and concentration differences) without Energy Expenditure. It is a selective process—a substance will only be transported across the membrane if a functional transport protein is available for it. Because facilitated diffusion involves proteins, the process, unlike passive diffusion, can reach saturation. The saturation plateau (Fig. 4.10) reflects a state in which all transport proteins for a given substance are saturated with substrate, and the transport rate reaches its maximum.

Facilitated diffusion mediates the Transmembrane Transport of numerous substances, including hydrophilic molecules such as CARBOHYDRATES, Amino Acids, NUCLEOTIDES, and various ions. Furthermore, the rate of this transport significantly exceeds that of passive diffusion (Fig. 4.10). In principle, substances and ions can cross the membrane via two distinct pathways: carriers and channels. Since Transmembrane Movement of carrier proteins themselves has not been observed in biomembranes, a “ping-pong” model has been proposed to describe carrier function. According to this mechanism, transport is coupled to Conformational Changes in the carrier Protein Structure induced by the binding of the transported substance (Fig. 4.11).

Fig. 4.10. Dependence of the rates of passive and facilitated diffusion on the concentration gradient of the transported substance.

Fig. 4.11. The “ping-pong” model of integral transport protein function.

Channel operation can be illustrated by the Acetylcholine Receptor. This integral protein is located primarily in the membranes of Skeletal Muscle neuromuscular junctions. It consists of five subunits of four different types and opens in response to the binding of acetylcholine (a neurotransmitter). Upon interaction with acetylcholine, the channel opens—a process driven by a conformational shift in the subunits—and allows specific ions (Na⁺, K⁺, Ca⁺⁺, and certain others) to pass through; it remains open for 1 ms before closing again. The selective movement of cations alters the transmembrane potential, triggering electrical excitation of the muscle cell, which leads to Muscle contraction. Structural Analysis of the acetylcholine receptor has revealed that the five protein subunits are embedded in the bilayer in a specific orientation: they are arranged around a central pore with a diameter of 3 nm, through which cations are transported. The channel's impermeability to anions and its threefold higher permeability to cations compared to uncharged molecules can be explained by Electrostatic Interactions arising from the presence of zwitterionic or negatively charged groups at the channel gate.

A distinct transport mechanism within The Scope of facilitated diffusion involves The Use of ionophores, the effects of which have been studied in Artificial Membranes. Ionophores are low-molecular-weight peptidic substances that selectively transport ions across membranes. They are categorized into channel-forming ionophores (such as gramicidin A and amphotericin B) and carrier ionophores (such as valinomycin, enniatins, and beauvericin).

Valinomycin is a depsipeptide antibiotic that Functions as an ionic “trap.” In nonpolar Solvents, the conformation of valinomycin resembles a bracelet, with an internal cavity precisely tailored to fit potassium ions. The outer surface of valinomycin is hydrophobic, enabling it to move freely within the lipid bilayer and transport ions across it.

A well-characterized example of a channel-forming ionophore is gramicidin A, a peptidic antibiotic consisting of 15 amino acids. Two gramicidin molecules can span the membrane as a double helix or by forming a HEAD-to-head dimer. In these Conformations, gramicidin A molecules create a hollow cylinder through which Metal Ions can travel.

An ionophore-mediated transport mechanism has not yet been identified in Introduction/36.html">Biological Membranes.

Active transport. This is the energy-dependent translocation of molecules or ions across a membrane against an electrochemical gradient, mediated by transport proteins. Thanks to active transport, living Cells maintain a potential difference—an electrical charge—across the membrane, with the interior of most studied cells being negatively charged relative to the external environment.

Active transport entails significant energy expenditure: some cells consume more than one-third of their total stored energy merely to generate ionic gradients across the membrane. This is essential for such vital processes as osmoregulation, the generation and transmission of nerve impulses, and the uptake of nutrients (such as sugars and amino acids) into cells.

Various active transport systems differ primarily in their energy source: ATP, ion gradients, phosphoenolpyruvate, or visible light. The best-characterized active transport system is the sodium-potassium pump (Na⁺/K⁺-ATPase), which operates in the plasma membranes of animal cells. This integral protein consists of two subunits: the larger one is a polypeptide featuring binding sites for sodium ions and ATP on the cytoplasmic surface, and for potassium ions on the outer surface; the smaller subunit is a glycoprotein. The operation of the (Na⁺/K⁺)-ATPase proceeds as follows: the Hydrolysis of a single ATP molecule pumps 3 Na⁺ ions out of the cell while importing 2 K⁺ ions, meaning that more positive ions are expelled than are brought in. This generates an excess negative charge on the inner face of the membrane, rendering the cell electrogenic. While pathways for the passive facilitated diffusion of sodium and potassium ions down their electrochemical gradients generally exist in membranes—allowing this transport to occur, albeit slowly—a living cell prevents the dissipation of these concentration gradients through the continuous activity of primary active transporters, such as the (Na⁺/K⁺)-ATPase. Thus, ion pumps involved in primary active transport generate Membrane Potential and establish an energy-storing electrochemical gradient.

Secondary active transporters utilize electrochemical gradients as the driving force for The transport of solutes. This process can be observed in intestinal epithelial cells. The building blocks generated by food Digestion IN THE intestine (such as Amino Acids and glucose) enter the bloodstream via diffusion across the membranes of Blood Vessels, a process facilitated by symport (cotransport) with sodium ions. Sodium ions tend to flow back into the cell in accordance with facilitated diffusion, effectively dragging nutrient molecules along with them. The membranes contain specific sugar and amino acid carriers that operate within an active transport system, accumulating these substances within the cell by extracting them even from highly dilute solutions—that is, against a chemical gradient. These same transport systems can also participate in facilitated diffusion if the substances are moving down their chemical gradient. In addition to the aforementioned symport of nutrients alongside re-entering sodium ions, counter-transport, or antiport, also exists. For instance, the erythrocyte anion exchanger 1 (band 3 protein) mediates the coupled, opposite-direction transport of Cl^- and HCO₃^- across THE RED BLOOD cell membrane.

In aerobic Bacteria, nutrient uptake is driven by symport with protons rather than Na⁺ ions. The best-characterized example of such a transporter is the lactose permease of Escherichia coli. This integral protein utilizes the proton electrochemical gradient established across the membrane by Oxidative Phosphorylation to drive lactose symport: for every proton returned to the cell, one lactose molecule is transported.

It is worth noting that ATPases are Enzymes that catalyze reversible reactions: during ATP hydrolysis, ions are transported against an electrochemical gradient, whereas the movement of ions down an electrochemical gradient through ATPase channels can drive the synthesis of ATP.

Endocytosis and Exocytosis. These mechanisms of transmembrane transport involve The formation of membrane invaginations and specialized membrane vesicles that facilitate the passage of large macromolecules and particles across the membrane. Endocytosis mediates the uptake of substances into the cell, whereas exocytosis drives their secretion. Cytosis is further subdivided into two types: pinocytosis and phagocytosis. Pinocytosis is the process by which proteins and other fluid-phase macromolecules are transported across the membrane. Phagocytosis involves the internalization of large particles, such as bacteria and Viruses. These transport modes are characteristic primarily of Eukaryotic cells; in animals, however, phagocytosis is restricted to specialized cells like macrophages. For many Protozoans, such as amoebas, phagocytosis serves as the primary mode of Nutrition.

A crucial feature of cytosis is the sequential formation and fusion of vesicles encapsulating the transported substance. The secreted or internalized molecules remain sequestered within these vesicles and do not mix with other macromolecules or cellular organelles. Through a mechanism not yet fully elucidated, each vesicle fuses exclusively with specific membrane structures (Fig. 4.12).

Cytosis is also underpinned by another hallmark property of biomembrane lipid layers—their capacity for aggregation, whereby small vesicles coalesce into larger ones or fuse with The Plasma Membrane. This mechanism relies on the structural universality of biomembranes involved in forming cellular organelles and protoplasts. A comparable phenomenon can be observed in foams, where soap bubbles—similarly composed of amphiphilic molecules (soaps, which are fatty acid salts)—tend to coalesce into larger structures. The aggregative capacity of membranes forms the basis for protoplast fusion, a widely used technique for transferring Genetic information.

The rate of cytosis is remarkably high. Studies have shown that within just one hour, Liver cells internalize a volume of fluid via endocytosis equal to at least 20% of their total volume, along with an amount of membrane material whose surface area is five times that of their plasma membrane. Like Other forms of transmembrane transport, cytosis requires that the transported substance be "recognized" by membrane components; in other words, selective membrane permeability to various compounds is evident here as well.

Fig. 4.12. Schematic representation of cytosis. Stages: 1-1 — a secretory vesicle buds off from the Golgi apparatus; 1-2 — the vesicle diffuses toward the plasma membrane and adheres to it; 1-3 — vesicle-membrane fusion and release of the vesicle contents into the extracellular environment; 2-1 — a particle is engulfed by the membrane; 2-2 — a phagosome pinches off from the membrane; 2-3 — the phagosome diffuses toward a lysosome; 2-4 — the phagosome adheres to the lysosome and fuses with it.