Читать книгу Handbook of Enology: Volume 1 - Pascal Ribéreau-Gayon - Страница 20

The plasma membrane is a highly selective barrier controlling exchanges between the living cell and its external environment. This organelle is essential to the life of the yeast.

Like all biological membranes, the yeast plasma membrane is principally made up of lipids and proteins. The plasma membrane of S. cerevisiae contains about 40% lipids and 50% proteins. Glucans and mannans are only present in small quantities (a few percent).

The lipids of the membrane are essentially phospholipids and sterols. They are amphiphilic molecules, i.e. possessing a hydrophilic and a hydrophobic part.

The three principal phospholipids (Figure 1.5) of the plasma membrane of yeast are phosphatidylethanolamine (PE), phosphatidylcholine (PC), and PI, which represent 70–85% of the total. Phosphatidylserine (PS) and diphosphatidylglycerol or cardiolipin (PG) are less prevalent. Free fatty acids and phosphatidic acid are frequently reported in plasma membrane analysis. They are probably extraction artifacts caused by the activity of certain lipid degradation enzymes.

The fatty acids of the membrane phospholipids contain an even number (14–24) of carbon atoms. The most abundant are C16 and C18 acids. They can be saturated, such as palmitic acid (C16) and stearic acid (C18), or unsaturated, as with oleic acid (C18, double bond in position 9), linoleic acid (C18, two double bonds in positions 9 and 12), and linolenic acid (C18, three double bonds in positions 9, 12, and 15). All membrane phospholipids share a common characteristic: they possess a polar or hydrophilic part made up of a phosphorylated alcohol and a nonpolar or hydrophobic part comprising two more‐or‐less parallel fatty acid chains. Their symbolic representation is shown in Figure 1.6. In an aqueous medium, the phospholipids spontaneously form bimolecular films or a lipid bilayer because of their amphiphilic nature. The lipid bilayers are cooperative but non‐covalent structures. They are maintained in place by mutually reinforced interactions: hydrophobic interactions and van der Waals forces between the hydrocarbon tails, and hydrostatic interactions and hydrogen bonds between the polar heads and water molecules. The examination of cross‐sections of yeast plasma membranes under an electron microscope reveals a classic lipid bilayer structure with a thickness of about 7.5 nm. The membrane surface appears sculpted with creases, especially during the stationary phase. However, the physiological meaning of this anatomical characteristic remains unknown. The plasma membrane also has a depression under the bud scar.

Ergosterol is the primary sterol of the yeast plasma membrane. In addition, 24(28)‐dehydroergosterol and lesser amounts of zymosterol are present (Figure 1.7). Sterols are exclusively produced in the mitochondria under aerobic conditions during the yeast log phase. As with phospholipids, membrane sterols are amphipathic. The hydrophilic part is composed of the hydroxyl group in the C3 position, while the rest of the molecule is hydrophobic, especially the flexible hydrocarbon tail.

The plasma membrane also contains numerous proteins or glycoproteins presenting a wide range of molecular weights (from 10,000 to 120,000). The available information indicates that the organization of the plasma membrane of a yeast cell resembles the fluid mosaic model. This model, proposed for biological membranes by Singer and Nicolson (1972), consists of two‐dimensional solutions of proteins and oriented lipids. Certain proteins penetrate the membrane; they are called integral proteins (Figure 1.6). They interact strongly with the nonpolar part of the lipid bilayer. The peripheral proteins are linked to the integral ones by hydrogen bonds. Their location is asymmetrical, at either the inner or the outer side of the plasma membrane. The molecules of proteins and membrane lipids, constantly in lateral motion, are capable of rapidly diffusing in the membrane.

Some of the yeast membrane proteins have been studied in greater depth. These include adenosine triphosphatase (ATPase), solute (sugars and amino acids) transport proteins, and enzymes involved in the production of glucans and chitin of the cell wall.

FIGURE 1.5 Yeast membrane phospholipids.

FIGURE 1.6 Diagram of a membrane lipid bilayer. The integral proteins (a) are strongly associated to the hydrocarbon region of the bilayer. The peripheral proteins (b) are linked to the integral proteins.

FIGURE 1.7 Principal yeast membrane sterols.

Yeast possesses three ATPases: one each in the mitochondria, the vacuole, and the plasma membrane. The plasma membrane ATPase is an integral protein with a molecular weight of around 100,000 Da. It catalyzes the hydrolysis of adenosine triphosphate (ATP), which furnishes the necessary energy for the active transport of solutes across the membrane. (Note: active transport moves a compound against the concentration gradient.) Simultaneously, the hydrolysis of ATP creates an efflux of protons toward the exterior of the cell.

The penetration of amino acids and sugars into the yeast activates membrane transport systems called permeases. General amino acid permease (GAP) contains three membrane proteins and ensures the transport of a number of neutral amino acids. The cultivation of yeasts in the presence of an easily assimilated nitrogen‐based nutrient such as ammonium represses this permease.

The fatty acid composition of the membrane and its sterol content control its fluidity. The hydrocarbon chains of fatty acids of the membrane phospholipid bilayer can be in a rigid and orderly state or in a relatively disorderly and fluid state. In the rigid state, all of the carbon bonds of the fatty acids are trans. In the fluid state, some of the bonds become cis. The transition from the rigid state to the fluid state takes place when the temperature rises beyond the melting point. This transition temperature depends on the length of the fatty acid chains and their degree of unsaturation. The straight hydrocarbon chains of the saturated fatty acids interact strongly. These interactions intensify with their length. The transition temperature therefore increases as the fatty acid chains become longer. The double bonds of the unsaturated fatty acids are generally cis, giving a curvature to the hydrocarbon chain (Figure 1.8). This curvature breaks the orderly stacking of the fatty acid chains and lowers the transition temperature. Like cholesterol in the cells of mammals, ergosterol is also a fundamental regulator of membrane fluidity in yeasts. Ergosterol is inserted in the bilayer perpendicularly to the plane of the membrane. Its hydroxyl group is bound by hydrogen bonds with the polar head of the phospholipid, and its hydrocarbon tail is inserted in the hydrophobic region of the bilayer. Thus, the membrane sterols insert themselves between the phospholipids. In this manner, they inhibit the crystallization of the fatty acid chains at low temperatures. Conversely, in reducing the movement of these same chains by steric encumbrance, they regulate any excessive membrane fluidity when the temperature rises.

FIGURE 1.8 Molecular models representing the three‐dimensional structure of stearic and oleic acids. The cis configuration of the double bond of oleic acid produces a curvature of the carbon chain.