Biochemistry SemV PaperC2 Unit1 Dr Rita Mahapatra Assistant

Biochemistry Sem-V Paper-C-2, Unit-1 Dr. Rita Mahapatra Assistant Professor Neotech College of applied Science and Research, Virod, Vadodara

Plant cell structure • The term cell Is derived from the Latin cella, meaning storeroom or chamber was given by Robert Hooke in 1665. • Plants are multicellular organisms composed of millions of cells with specialized functions. At maturity, such specialized cells may differ greatly from one another in their structures. • However, all plant cells have the same basic eukaryotic organization (Fig 1): They contain § 1: Cytoplasm (Chloroplast, mitochondria etc. ) § 2: Nucleus § 3: Cell membrane § 4: Cell wall § 5: Chromosomes § 6: Plasmodesmata § 7: Filamentous cytoskeleton

Fig 1. Plant cell structure

Cell membrane • The cytoplasm and the nucleus and other parts of the cell are enclosed within the cell membrane or plasma membrane which separates cell from one another and also the cell from the surrounding medium. • The membrane is porous and allows the movement of substances or materials both inward and outward. The yellow outline in this diagram is the cell membrane.

Cell Wall • The cell wall is the most characteristic feature of a plant cell • The cell wall is always non-living but is formed and maintained by the living organism • Its primary function is to provide protection to the contents of cell • Due to semi-rigid nature, the cell walls are responsible for giving • shapes to different kinds of cells during cell differentiation of • Tissues • In multicellular and woody plants of cell wall is differentiated into • three parts i. e. , the middle lamella, the primary wall and • secondary wall. • The cell wall is a very tough, flexible and sometimes fairly rigid layer that surrounds plant cells. It surrounds the cell membrane and provides these cells with structural support and protection. • In addition the cell wall is acting as a filtering mechanism. A major function of the cell wall is to act as a pressure vessel, preventing over-expansion when water enters the cell. The green outline in this diagram is the cell wall: -

The middle lamella • It is a common structure between adjacent cells and therefore, binds them • with each other. • It is an amorphous layer and is • composed of calcium and magnesium pectate. • The middle lamella remains unlignified • in case of softer living tissues namely • Parenchyma, collenchyma and arenchyma, but in woody tissues Sclerenchyma it becomes highly lignified

The primary cell wall • Consists of cellulose (45%), hemicellulose (25%), pectins • (35%) and structural proteins (upto 8%) on the basis of dry • weight • The primary wall is thin and elastic • It is capable of growth and expansion • The backbone of the primary wall is formed by the cellulose • fibrils. • The matrix is composed of hemicellulose, pectin, gums, • tannins, resins, silica, waxes etc. and small structured proteins

Secondary cell wall • The 20 wall is very thick (lignin), rigid and inelastic and consists of three layers known as S 1 (outer), S 2 (middle) and S 3 (inner) • The microfibrils in these layers run parallel to each other but the directions are different in three layers. • The microfibrils are transversely arranged in the S 3 and are at an angle of • 10 -200 to the longitudinal axis in S 2 and are at the angle of 500 in S 1 • The lignin is formed from three different phenyl propanoid alcohols: coniferyl, coumaryl and sinapyl alcohols. • Lignin is covalently bonded to cellulose and other polysaccharides of cell wall.

Difference between the primary and secondary cell wall

Functions of cell wall • They determine the morphology, growth, and development of plant cells. • They protect the protoplasm from invasion by viral, bacterial and fungal pathogens. • They are rigid structures and thus help the plant in withstanding the gravitational forces. • They are involved in the transport of materials and metabolites into and out of cell. • They withstand the turgor pressure which develops within the cells due to high osmotic pressure.

Nucleus • The nucleus is double membrane-enclosed organelle found in eukaryotic cells. The nucleus is generally spherical and located in the centre of the cell. The nucleus contains thread like structures called chromosomes which carry genes. Gene is a unit of inheritance in living organisms. • The function of the nucleus is to maintain the integrity of these genes and to control the activities of the cell by regulating gene expression—the nucleus is, therefore, the control center of the cell. • Nuclei contain a densely granular region, called the nucleolus (plural nucleoli), that is the site of ribosome synthesis. • A specific amino acid sequence called the nuclear localization signal is required for a protein to gain entry into the nucleus. • The main structures making up the nucleus are the nuclear envelope, a double membrane that encloses the entire organelle and isolates its contents from the cellular cytoplasm, the nucleolus and nuclear pores. • Nuclear pores regulate the transport of molecules across the envelope. • The nucleolus is a smaller spherical body in the nucleus.

Cytoplasm • The jelly- like substance present between the cell membrane and nucleus is known as the cytoplasm. The cytoplasm is about 80% water and usually colorless. Various other components, or organelles, of cells are present in the cytoplasm. These are : • Mitochondria • Chloroplast • Golgi bodies • Ribosomes • Endoplasmic reticulum • Lysosomes • Vacuole • Peroxisomes etc.

Chloroplast • Chloroplast belong to another group of double membrane–enclosed organelles called plastids. • Chloroplast membranes are rich in glycosylglycerides. • Chloroplast membranes contain chlorophyll and it’s associated proteins and are the sites of photosynthesis. • In addition to their inner and outer envelope membranes, chloroplasts possess a third system of membranes called thylakoids. A stack of hylakoids forms a granum (plural grana) (Fig. 2). Proteins and pigments (chlorophylls and carotenoids) that function in the photochemical events of photosynthesis are embedded in the thylakoid membrane. • The fluid compartment surrounding the thylakoids, called the stroma, is analogous to the matrix of the mitochondrion. • Adjacent grana are connected by unstacked membranes called stroma lamellae (singular lamella). Plastids that contain high concentrations of carotenoid pigments rather than chlorophyll are called chromoplasts. They are one of the causes of the yellow, orange, or red colors of many fruits and flowers, as well as of autumn leaves •

Fig. 2. Chloroplast

Cont. • Non pigmented plastids are called leucoplasts. The most important type of leucoplast is the amyloplast, a starchstoring plastid. • Amyloplasts are abundant in storage tissues of the shoot and root, and in seeds. Specialized amyloplasts in the root cap also serve as gravity sensors that direct root growth downward into the soil. • Chlorophylls are pigments in chloroplast absorb red and blue light and reflect green and yellow light to excite electron (e-) – plant appears green. • Plant absorbs CO 2, H 2 O and O 2 as raw material of photosynthesis. • Carbon dioxide + water → Sugar + Oxygen + • Water + Sunlight • 6 CO 2 + 6 H 2 O → C 6 H 12 O 6 + 6 O 2

Mitochondria • Mitochondria (singular mitochondrion) are the cellular sites of respiration, a process in which the energy released from sugar metabolism is used for the synthesis of ATP (adenosine triphosphate) from ADP (adenosine diphosphate) and inorganic phosphate (Pi). • Mitochondria can vary in shape from spherical to tubular, but they all have a mooth outer membrane and a highly convoluted inner membrane (Fig. 3). • The infoldings of the inner membrane are called cristae (singular crista). • The compartment enclosed by the inner membrane, the mitochondrial matrix, contains the enzymes of the pathway of intermediary metabolism called the Krebs cycle. • In contrast to the mitochondrial outer membrane and all other membranes in the cell, the inner membrane of a mitochondrion is almost 70% protein and contains some phospholipids that are unique to the organelle (e. g. , cardiolipin). • The proteins in and on the inner membrane have special enzymatic and transport capacities. • The transmembrane enzyme ATP synthase is coupled to the phosphorylation of ADP to produce ATP.

Fig. 3 Mitochondria

Mitochondria and Chloroplasts Are Semiautonomous Organelles • Both mitochondria and chloroplasts contain their own DNA and protein-synthesizing machinery (70 S ribosomes, transfer RNAs, and other components) and are believed to have evolved from endosymbiotic bacteria. • Both plastids and mitochondria divide by fission, and mitochondria can also undergo extensive fusion to form elongated structures or networks. • The DNA of Mitochondria 200 Kb and chloroplast 145 Kb, these organelles is in the form of circular chromosomes, similar to those of bacteria and very different from the linear chromosomes in the nucleus. • These DNA circles are localized in specific regions of the mitochondrial matrix or plastid stroma called nucleoids. • DNA replication in both mitochondria and chloroplasts is independent of DNA replication in the nucleus.

Peroxisomes • Peroxisomes are found in all eukaryotic organisms, and in plants they are present in photosynthetic cells Peroxisomes function both in the removal of hydrogens from organic substrates, consuming O 2 in the process, according to the following reaction: • RH 2 + O 2 → R + H 2 O 2 (where R is the organic substrate) • The potentially harmful peroxide produced in these reactions is broken down in peroxisomes by the enzyme catalase, according to the following reaction: • H 2 O 2 → H 2 O + 1⁄2 O 2 • Although some oxygen is regenerated during the catalase • reaction, there is a net consumption of oxygen overall.

Glyoxysome • Another type of microbody, the glyoxysome, is present in oilstoring seeds. Glyoxysomes contain the glyoxylate cycle enzymes, which help convert stored fatty acids into sugars that can be translocated throughout the young plant to provide energy for growth • Because both types of microbodies (peroxisome & glyoxysome) carry out oxidative reactions, it has been suggested they may have evolved from primitive respiratory organelles that were superseded by mitochondria.

Vacuoles • Mature living plant cells contain large, water-filled central vacuoles that can occupy 80 to 90% of the total volume of the cell. • Each vacuole is surrounded by a vacuolar membrane, or tonoplast. Many cells also have cytoplasmic strands that run through the vacuole, but each transvacuolar strand is surrounded by the tonoplast. • The vacuole contains water and dissolved inorganic ions, organic acids, ugars, enzymes, and a variety of secondary metabolites, which often play roles in plant defense. • Active solute accumulation provides the osmotic driving force for water uptake by the vacuole, which is required for plant cell enlargement. • The turgor pressure generated by this water uptake provides the structural rigidity needed to keep herbaceous plants upright, since they lack the lignified support tissues of woody plants.

• Vacuoles contain many hydrolytic enzymes, such as proteases, ribonucleases, glycosidases, and phosphatases, as well as peroxidases and a variety of secondary metabolites, which often play roles in plant defense. Active solute accumulation provides the osmotic driving force for water uptake by the vacuole, which is required for plant cell nlargement. • The turgor pressure generated by this water uptake provides the structural rigidity needed to keep herbaceous plants upright, since they lack the lignified support tissues of woody plants. • Specialized protein-storing vacuoles, called protein bodies, are abundant in seeds. During germination the storage proteins in the protein bodies are hydrolyzed to amino acids and exported to the cytosol for use in protein synthesis. • The hydrolytic enzymes are stored in specialized lytic vacuoles, which fuse with the protein bodies to initiate the breakdown process

Plasmodesmata • Plasmodesmata are microscopic channels which allow molecules to travel between plant cells. Unlike animal cells, every plant cell is surrounded by a cell wall. Neighboring plant cells are therefore separated by a pair of cell walls, forming an extracellular domain known as the apoplast. • Although cell walls allow small soluble proteins and other solutes to pass through them, Plasmodesmata enable direct, regulated, symplastic intercellular transport of substances between cells. Plasmodesmata (singular plasmodesma) are tubular extensions of the plasma membrane, 40 to 50 nm in diameter, that traverse the cell wall and connect the cytoplasms of adjacent cells. Because most plant cells are interconnected in this way, their cytoplasms form a continuum referred to as the symplast. Intercellular transport of solutes through plasmodesmata is thus called symplastic transport

Filamentous cytoskeleton • The cytoskeletal components—microtubules, microfilaments, and intermediate filaments—participate in a variety of processes involving intracellular movements, such as mitosis, cytoplasmic streaming, secretory vesicle transport cell plate formation, and cellulose microfibril deposition. • The process by which cells reproduce is called the cell cycle. • The filamentous cytoskeleton is a network of fibers composed of proteins contained within a cell's cytoplasm. It is a dynamic structure, parts of which are constantly destroyed, renewed or newly constructed. • Here is a multitude of functions the cytoskeleton can perform: It gives the cell shape and mechanical resistance to deformation, it stabilizes entire tissues, it can actively contract, hereby deforming the cell and the cell's environment and allowing cells to migrate, it divides chromosomes. • It is involved in the division of a mother cell into two daughter cells etc. The functions which this cytoskeleton can perform depend on the type of cell and the organism.

Functions of Water in Plants • Water is important in the life of plants because it makes up the matrix and medium in which most biochemical processes essential for life take place. • It has been said that the study of plant physiology is, for the most part, the study of plant water relations. This is understandable, given the central role water plays in a large number of plant processes. • Among the many functions of water in plants are the following: · it serves as a medium (and sometimes substrate) for biochemical reactions in cells, since many enzymes are dissolved in the cell water · structural support – water provides the “turgor pressure” that gives many cells their shape; thus, many tissues will lose their structure and wilt when water availability is inadequate. • cell enlargement – turgor pressure provides the physical force needed to expand cells during growth. Transport of solutes between organs, via the xylem and phloem vessels evaporative cooling of leaves during transpiration. • Next to light, water availability is probably the single most important environmental factor affecting plant growth. Accordingly, plants have evolved with complex physiological strategies for regulating water use, including, but not limited to, minutes timescale regulation of stomatal apertures in response to sudden changes in environmental conditions. In cropping situations • Worldwide, water deficits constitute the single largest cause of crop failure.

Diffusion • • The net, random movement of individual molecules from one area to another. The molecules move from [hi] → [low], following a concentration gradient. Another way of stating this is that the molecules move from an area of high free energy (higher concentration) to one of low free energy (lower concentration). The net movement stops when a dynamic equilibrium is achieved.

• Some studies indicated that diffusion directly across the lipid bilayer was not sufficient to account for observed rates of water movement across membranes, but the evidence in support of microscopic pores was not compelling. • This uncertainty was put to rest with the recent discovery of aquaporins. Aquaporins are integral membrane proteins that form water-selective channels across the membrane. Because water diffuses faster through such channels than through a lipid bilayer, aquaporins facilitate water movement into plant cells. Water can cross plant membranes by diffusion of individual water molecules through the embrane bilayer, as shown on the left, and by microscopic bulk flow of water molecules through a waterelective pore formed by integral membrane proteins such as aquaporins as shown in the diagram

Osmosis The spontaneous net movement of solvent molecules through a partially permeable membrane into a region of higher solute concentration, in the direction that tends to equalize the solute concentrations on the two sides –dynamic equilibrium. • • • Water potential is a measure of the energy state of water. Determines the direction and movement of water. Unit for water potential Mega. Pascal Mpa Ψ pure water at 1 atm = 0 Mpa Ψ=Ψs + Ψp Ψ= water potential Ψs=solute potential (osmotic potential) –Always negative Ψp=(pressure potential) Water molecules move from higher water potential to lower water potential

ABSORPTION OF WATER • Water in the soil is mostly and abundantly, under normal conditions, is available in the form of Capillary water. Water plays a crucial role in the life of the plant. For every gram of organic matter made by the plant, approximately 500 g of water is absorbed by the roots, transported through the plant body and lost to the atmosphere. • In the soil the space in between soil particle forms a network of spaces, which normally is filled with water. • The water that is present in such spaces is called capillary water. • Land plants absorbs water by (1) An extensive root system to extract water from the soil; (2) A low-resistance pathway through the xylem vessel elements and tracheids to bring water to the leaves; (3) A hydrophobic cuticle covering the surfaces of the plant to reduce evaporation; (4) Microscopic stomata on the leaf surface to allow gas exchange; and (5) Guard cells to regulate the diameter (and diffusional resistance) of the stomatal aperture.

• In plants, following two pathways are involved in the water movement. They are • (1) Apoplastic pathway • (2) Symplastic pathway • (3) Transmembrane pathway • 1. Apoplastic pathway : The apoplastic movement of water in plants occurs exclusively through the cell wall without crossing any membranes. The cortex receive majority of water through apoplastic way as loosely bound cortical cells do not offer any resistance. But the movement of water in root beyond cortex apoplastic pathway is blocked by casparian strip present in the endodermis.

2. Symplastic pathway • The movement of water from one cell to other cell through the plasmodesmata is called the symplastic pathway of water movement. This pathway comprises the network of cytoplasm of all cells inter-connected by plasmodermata.

3. The transmembrane pathway is the route followed by water that sequentially enters a cell on one side, exits the cell on the other side, enters the next in the series, and so on. In this pathway, water crosses at least two membranes for each cell in its path (the plasma membrane on entering and on exiting). • Transport across the tonoplast may also be involved.

• • • Translocation and conduction of water and minerals Some aspects of phloem translocation have been well established by extensive research over many years. These include the following: The pathway of translocation- Sugars and other organic materials are conducted throughout the plant in the phloem, specifically in cells called sieve elements. Sieve elements display a variety of structural adaptations that make them well suited for transport. Patterns of translocation. Materials are translocated in the phloem from sources (areas of photosynthate supply) to sinks (areas of metabolism or storage of photosynthate). Sources are usually mature leaves. Sinks include organs such as roots and immature leaves and fruits. Materials translocated in the phloem. The translocated solutes are mainly carbohydrates, and sucrose is the most commonly translocated sugar. Phloem sap also contains other organic molecules, such as amino acids, proteins, and plant hormones, as well as inorganic ions.

• Rates of movement in the phloem are quite rapid, well in excess of rates of diffusion. Velocities average 1 m h– 1, and mass transfer ratesrange from 1 to 15 g h– 1 cm– 2 of sieve elements. Other aspects of phloem translocation require further investigation, and most of these are being studied intensively at the present time. These aspects include the following: • Phloem loading and unloading. Transport of sugars into and out of the sieve elements is called sieve element loading and unloading, respectively. • Mechanism of translocation. • Photosynthate allocation and partitioning.

Guttation • Guttation is the appearance of drops of xylem sap on the tips or edges of leaves of some vascular plants, such as grasses. • Guttation is not to be confused with dew, which condenses from the atmosphere onto the plant surface Secretion of water on to the surface of leaves through specialized pores, or hydathodes. • The main cause of guttation in plants is root pressure, during night when root pressure is high sometimes den due to this pressure watery drops ooze out with the assistance of special structures which help in guttation called the hydathodes.

Transpiration • During the plant’s lifetime, water equivalent to 100 times the fresh weight of the plant may be lost through the leaf surfaces. Such water loss is called transpiration. • Transpiration is an important means of dissipating the heat input from sunlight. Heat dissipates because the water molecules that escape into the atmosphere have higher than- average energy, which breaks the bonds holding them in the liquid. • For a typical leaf, nearly half of the net heat input from sunlight is dissipated by transpiration. • Latent heat of vaporization is the energy needed to separate molecules from the liquid phase and move them into the gas phase at constant temperature—a process that occurs during transpiration. • For water at 25°C, the heat of vaporization is 44 k. J mol– 1—the highest value known for any liquid. Most of this energy is used to break hydrogen bonds between water molecules.

1. Stomatal transpiration: Transpiration that occurs through stomata called stomatal transpiration. This type of transpiration only occurs in its presence of sunlight (in daytime). • Because stomata open in the present of sunlight and close in the darkness. • In this method plants give out 80 -90% water in the form of vapor.

2. Cuticular transpiration: • Transpiration that occurs through the cuticle or cracks of thin cuticle layer of leaves and stems is said to be cuticular transpiration. • This is a day-night process. In this process, 5 -10% water is given out in the form of vapor.

3. Lenticular transpiration: • Sometimes transpiration occurs through lenticels, the small opening in the corky tissue covering stems and twigs, and this type of transpiration is said to be the lenticular transpiration. In this process, only 0. 1% water is given off of the forms of vapor.

• Transpiration is a process similar to evaporation. • It is a part of the water cycle, and it is the loss of water vapor from parts of plants (similar to sweating), especially in leaves but also in stems, flowers and roots. • Leaf surfaces are dotted with openings which are collectively called stomata, and in most plants they are more numerous on the undersides of the foliage. • The stomata are bordered by guard cells that open and close the pore. • Leaf transpiration occurs through stomata. • An Increase in Guard Cell Turgor Pressure Opens the Stomata • Guard cells function as multisensory hydraulic valves. environmental factors such as light intensity and quality, temperature, relative humidity, and intracellular CO 2 concentrations are sensed by guard cells, and these signals are integrated into well-defined stomatal responses. If leaves kept in the dark are illuminated, the light stimulus is perceived by the guard cells as an opening signal, triggering a series of responses that result in opening of the stomatal pore.

Guard cell • In guard cells the microfibril organization is different. • Kidney-shaped guard cells have cellulose microfibrils fanning out radially from the pore (Fig. A). Thus the cell girth is reinforced like a steel-belted radial tire, and the guard cells curve outward during stomatal opening In grasses, the dumbbell-shaped guard cells function like beams with inflatable ends. • As the bulbous ends of the cells increase in volume and swell, the beams are separated from each other and the slit between them widens (Fig. B).

Mechanism of Transpiration • The Cohesion–Tension Theory Explains Water. Transport in the Xylem. In theory, the pressure gradients needed to move water through the xylem could result from the generation of positive pressures at the base of the plant or negative pressures at the top of the plant. • It is mentioned previously that some roots can develop positive hydrostatic pressure in their xylem—the so-called root pressure. • However, root pressure is typically less than 0. 1 MPa and disappears when the transpiration rate is high, so it is clearly inadequate to move water up a tall tree. • Instead, the water at the top of a tree develops a large tension (a negative hydrostatic pressure), and this tension pulls water through the xylem. This mechanism, first proposed toward the end of the nineteenth century, is called the cohesion–tension theory of sap ascent because it requires the cohesive properties of water to sustain large tensions in the xylem water columns.

Root pressure theory • Root pressure is most likely to occur when soil water potentials are high and transpiration rates are low. • When transpiration rates are high, water is taken up so rapidly into the leaves and lost to the atmosphere that a positive pressure never develops in the xylem. • Plants that develop root pressure frequently produce liquid droplets on the edges of their leaves, a phenomenon known as guttation.

Water balance and Stress • Water in the plant can be considered a continuous hydraulic system, connecting the water in the soil with the water vapor in the atmosphere. • Transpiration is regulated principally by the guard cells, which regulate the stomatal pore size to meet the photosynthetic demand for CO 2 uptake while minimizing water loss to the atmosphere. • Water evaporation from the cell walls of the leaf mesophyll cells generates large negative pressures (or tensions) in the apoplastic water. These negative pressures are transmitted to the xylem, and they pull water through the long xylem conduition. • Although aspects of the cohesion–tension theory of sap ascent are intermittently debated, an overwhelming body of evidence supports the idea that water transport in the xylem is driven by pressure gradients. • When transpiration is high, negative pressures in the xylem water may cause cavitation (embolisms) in the xylem. • Such embolisms can block water transport and lead to severe water deficits in the leaf. • Water deficits are commonplace in plants, necessitating a host of adaptive responses that modify the physiology and development of plants.

• Water deficit can be defined as any water content of a tissue or cell that is below the highest water content exhibited at the most hydrated state. • When water deficit develops slowly enough to allow changes in developmental processes, water stress has several effects on growth, one of which is a limitation in leaf expansion. Leaf area is important because photosynthesis is usually proportional to it. However, rapid leaf expansion can adversely affect water availability. • Adaptation and acclimation to environmental stresses result from integrated events occurring at all levels of organization, from the anatomical and morphological level to the cellular, biochemical, and molecular level. For example, the wilting of leaves in response to water deficit reduces both water loss from the leaf and exposure to incident light, thereby reducing heat stress on leaves. • At the biochemical level, plants alter metabolism in various ways to accommodate environmental stresses, including producing osmoregulatory compounds such as proline and glycine betaine to adapt and acclimate to water deficit, salinity, chilling and freezing, heat, and oxygen deficiency in the root biosphere.

References • Plant Physiology, 3 rd ed, Lincoln Taiz and Eduardo Zeiger, Publisher: Sinauer Associates; 3 edition (2002). • Fundamental of biochemistry by D Voet, J. GVoet and C. W Pratt, John Wiley & Sons, Inc. , New York, 2 nd edition, 2005 • Principles of Biochemistry by Albert Lehninger, W. H. Freeman & Company; 3 rd edition (February 2000), ISBN-10: 1572591536 • Botany by A. K Nanda