Proteins are very important molecules in our cells. They are involved in virtually all cell functions.

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2 Proteins are very important molecules in our cells. They are involved in virtually all cell functions. Proteins do everything in the living cells. All functions of the living organisms are related with proteins. Each protein within the body has a specific function. Some proteins are involved in structural support, while others are involved in bodily movement, or in defense against germs. Proteins vary in structure as well as function. They are constructed from a set of 20 amino acids and have distinct three-dimensional shapes.

3 Proteins are responsible for many different functions in the living cell. It is possible to classify proteins on the basis of their functions. Very often, proteins can carry few functions and such proteins can be placed into different groups, but despite this, it is possible to assign main group for each protein. Enzymes - catalytic activity and function Transport Proteins - bind & carry ligands Storage Proteins - ovalbumin, gluten, casein, ferretin Contractile (Motor): can contract, change shape, elements of cytoskeleton (actin, myosin, tubulin)

4 Structural (Support): collagen of tendons & cartilage, elastin of ligaments (tropoelastin), keratin of hair, feathers, & nails, fibroin of silk & webs Defensive (Protect): antibodies (IgG), fibrinogen & thrombin, snake venoms, bacterial toxins Regulatory (Signal): regulate metabolic processes, hormones, transcription factors & enhancers, growth factor proteins Receptors (detect stimuli): light & rhodopsin, membrane receptor proteins and acetylcholine or insulin.

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6 Enzymes - are proteins that facilitate biochemical reactions. They are often referred to as catalysts because they speed up chemical reactions. This group of proteins probably is the biggest and most important group of the proteins Examples include the enzymes DNA- and RNApolymerases, dehydrogenases, lactase and pepsin. Lactase breaks down the sugar lactose found in milk. Pepsin is a digestive enzyme that works in the stomach to break down proteins in food.

7 Catalyst: inorganic or organic substance which speeds up the rate of a chemical reaction without entering the reaction itself. Enzymes: organic catalysts made of protein. Like all catalysts, enzymes work by lowering the activation energy (E a or ΔG ) for a reaction, thus dramatically increasing the rate of the reaction. Most enzyme reaction rates are millions of times faster than those of comparable un-catalyzed reactions. most enzyme names end in -ase

8 Many enzymes catalyze reactions without help, but some require an additional non-protein component called a cofactor. Co-factors may be inorganic ions such as Fe 2+, Mg 2+, Mn 2+, or Zn 2+, or consist of organic or metalloorganic molecules knowns as co-enzymes. Enzyme activity can be affected by other molecules. Inhibitors are molecules that decrease enzyme activity; activators are molecules that increase activity. Many drugs and poisons are enzyme inhibitors. Activity is also affected by temperature, chemical environment (e.g., ph), and the concentration of substrate.

9 Some enzymes are used commercially, for example, in the synthesis of antibiotics. In addition, some household products use enzymes to speed up biochemical reactions (e.g., enzymes in biological washing powders break down protein or fat stains on clothes; enzymes in meat tenderizers break down proteins, making the meat easier to chew).

10 substrate: molecules upon which an enzyme acts. The enzyme is shaped so that it can only lock up with a specific substrate molecule. Substrae Sucrose enzyme Sucrase product Fructose + Glucose

11 Enzymes are very specific, both the enzyme and the substrate possess specific complementary geometric shapes that fit exactly into one another. This is often referred to as "the lock and key" model. However, while this model explains enzyme specificity, it fails to explain the stabilization of the transition state that enzymes achieve. The "lock and key" model has proven inaccurate, and the induced fit model is the most currently accepted enzyme-substrate-coenzyme figure.

12 Lock and Key Theory Each enzyme is specific for one and ONLY one substrate (one lock - one key) active site: part of the enzyme that fits with the substrate Note that the active site has a specific fit for this particular substrate and no other. This theory has some weaknesses, but it explains many basic things about enzyme function. how_enzymes_work.ht ml

13 Enzymes are classified according to the reactions they catalyze. The six classes are: Oxidoreductases Transferases Hydrolases Lyases Isomerases Ligases Examples: Alcohol dehydrogenase: an oxidoreductase converting alcohols to aldehydes/ ketones. Aminotransferases: transferases catalyzing the amino acid degradation by removing amino groups.

14 Glucose-6-phosphatase: a hydrolase that removes the phosphate group from glucose-6-phosphate, leaving glucose and H 3 PO 4. Pyruvate decarboxylase: a lyase that removes CO 2 from pyruvate. Ribulose phosphate epimerase: an isomerase that catalyzes the interconversion of ribulose-5-phosphate and xylulose-5- phosphate.

15 ph: the optimum (best) in most living things is close to 7 (neutral). High or low ph levels usually slow enzyme activity

16 Temperature: strongly influences enzyme activity optimum (best) temperature for maximum enzyme function is usually about C. reactions proceed slowly below optimal temperatures above 45 C. most enzymes are denatured (change in their shape so the enzyme active site no longer fits with the substrate and the enzyme can't function)

17 Concentrations of Enzyme and Substrate When there is a fixed amount of enzyme and an excess of substrate molecules the rate of reaction will increase to a point and then level off. This leveling off occurs because all of the enzyme is used up and the excess substrate has nothing to combine with. If more enzyme is available than substrate, a similar reaction rate increase and leveling off will occur. The excess enzyme will eventually run out of substrate molecules to react with.

18 Enzymes serve a wide variety of functions inside living organisms. They are indispensable for signal transduction and cell regulation, often via kinases and phosphatases. They also generate movement, with myosin hydrolysing ATP to generate muscle contraction and also moving cargo around the cell as part of the cytoskeleton. Other ATPases in the cell membrane are ion pumps involved in active transport. Enzymes are also involved in more exotic functions, such as luciferase generating light in fireflies.

19 Viruses can also contain enzymes for infecting cells, such as the HIV integrase and reverse transcriptase, or for viral release from cells, like the influenza virus neuraminidase. An important function of enzymes is in the digestive systems of animals. Enzymes such as amylases and proteases break down large molecules (starch or proteins, respectively) into smaller ones, so they can be absorbed by the intestines. Starch molecules, for example, are too large to be absorbed from the intestine, but enzymes hydrolyse the starch chains into smaller molecules such as maltose and eventually glucose, which can then be absorbed. Different enzymes digest different food substances. In ruminants which have a herbivorous diets, microorganisms in the gut produce another enzyme, cellulase to break down the cellulose cell walls of plant fiber.

20 Antibodies are substances made by the body's immune system in response to bacteria, viruses, fungus, animal dander, or cancer cells. Antibodies attach to the foreign substances so the immune system can destroy them. Antibodies are specific to each type of foreign substance. For example, antibodies made in response to a tuberculosis infection attach only to tuberculosis bacteria. Antibodies may be made against your own tissues. This is called an autoimmune disease.

21 If your immune system makes low levels of antibodies, you may have a higher chance of developing repeated infections. You can be born with an immune system that makes low levels of antibodies, or your system may make low levels of antibodies in response to certain diseases, such as cancer. Antibodies are produced by a kind of white blood cell called a B cell. Antibodies occur in two forms: membrane-bound (attached to the surface of a B cell) Soluble (secreted by specific B cells called plasma cells)

22 Stimulating B Cell When a T lymphocyte "sees" the same peptide on the macrophage and on the B cell, the T cell stimulates the B cell to turn on antibody production. Antibody Production The stimulated B cell undergoes repeated cell divisions, enlargement and differentiation to form a clone of antibody secreting plasma cells. Hence. through specific antigen recognition of the invader, clonal expansion and B cell differentiation you acquire an effective number of plasma cells all secreting the same needed antibody. That antibody then binds to the bacteria making them easier to ingest by white cells. Antibody combined with a plasma component called "complement" may also kill the bacteria directly.

23 The general structure of all antibodies is very similar Each antibody consists of four polypeptides two heavy chains and two light chains held together by disulfide bonds joined to form a "Y" shaped molecule. The amino acid sequence in the tips of the "Y" varies greatly among different antibodies. This variable region, composed of amino acids, give the antibody its specificity for binding antigen.

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25 A small region at the tip of the protein is extremely variable, allowing millions of antibodies with slightly different tip structures to exist. This region is known as the hypervariable region. Each of these variants can bind to a different target, known as an antigen. This huge diversity of antibodies allows the immune system to recognize an equally wide diversity of antigens.

26 Neutralization of toxins Immobilization of microorganisms Neutralization of viral activity Agglutination Binding soluble antigen (precipitation) Activating complement

27 Activation of complement Antibodies that bind to surface antigens on, for example a bacterium, attract the first component of the complement cascade with their Fc* region and initiate activation of the "classical" complement system. This results in the killing of bacteria in two ways. First, the binding of the antibody and complement molecules marks the microbe for ingestion by phagocytes in a process called opsonization; these phagocytes are attracted by certain complement molecules generated in the complement cascade. Secondly, some complement system components form a membrane attack complex to assist antibodies to kill the bacterium directly. * The base of the Y plays a role in modulating immune cell activity. This region is called the Fc (Fragment, crystallizable) region, and is composed of two heavy chains that contribute two or three constant domains depending on the class of the antibody

28 Activation of effector cells To combat pathogens that replicate outside cells, antibodies bind to pathogens to link them together, causing them to agglutinate. Since an antibody has at least two paratopes it can bind more than one antigen by binding identical epitopes carried on the surfaces of these antigens. By coating the pathogen, antibodies stimulate effector functions against the pathogen in cells that recognize their Fc region. Those cells which recognize coated pathogens have Fc receptors which, as the name suggests, interacts with the Fc region of IgA, IgG, and IgE antibodies.

29 The engagement of a particular antibody with the Fc receptor on a particular cell triggers an effector function of that cell; phagocytes will phagocytose, mast cells and neutrophils will degranulate, natural killer cells will release cytokines and cytotoxic molecules; that will ultimately result in destruction of the invading microbe. The Fc receptors are isotype-specific, which gives greater flexibility to the immune system, invoking only the appropriate immune mechanisms for distinct pathogens.

30 The immunoglobulins can be divided into five different classes, based on differences in the amino acid sequences in the constant region of the heavy chains. IgA - Alpha heavy chains: IgA antibodies are found in areas of the body such the nose, breathing passages, digestive tract, ears, eyes, and vagina. IgA antibodies protect body surfaces that are exposed to outside foreign substances. This type of antibody is also found in saliva, tears, and blood. About 10% to 15% of the antibodies present in the body are IgA antibodies. A small number of people do not make IgA antibodies.

31 IgG - Gamma heavy chains: IgG antibodies are found in all body fluids. They are the smallest but most common antibody (75% to 80%) of all the antibodies in the body. IgG antibodies are very important in fighting bacterial and viral infections. IgG antibodies are the only type of antibody that can cross the placenta in a pregnant woman to help protect her baby. IgM - Mu heavy chains: IgM antibodies are the largest antibody. They are found in blood and lymph fluid and are the first type of antibody made in response to an infection. They also cause other immune system cells to destroy foreign substances. IgM antibodies are about 5% to 10% of all the antibodies in the body.

32 IgE - Epsilon heavy chains: IgE antibodies are found in the lungs, skin, and mucous membranes. They cause the body to react against foreign substances such as pollen, fungus spores, and animal dander. They may occur in allergic reactions to milk, some medicines, and some poisons. IgE antibody levels are often high in people with allergies. IgD - Delta heavy chains: IgD antibodies are found in small amounts in the tissues that line the belly or chest. How they work is not clear.

33 Immunoglobulin Types: Immunoglobulins can also be classified by the type of light chain that they have. Light chain types are based on differences in the amino acid sequence in the constant region of the light chain. These differences are detected by serological means. 1. Kappa light chains 2. Lambda light chains

34 Storage proteins are biological reserves of metal ions and amino acids, used by organisms. They are found in plant seeds, egg whites, and milk. Ferritin is an example of a storage protein that stores iron. Iron is a component of heme, which is contained in the transport protein hemoglobin and in cytochromes. Some storage proteins store amino acids. Storage proteins' amino acids are used in embryonic development of animals or plants. Two amino acid storage proteins in animals are casein and ovalbumin. Seeds, particularly of leguminous plants, contain high concentrations of storage proteins. Up to 25 percent of the dry weight of the seed can be composed of storage proteins.

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36 Ovalbumin: Ovalbumin is a glycoprotein with molecular weight of 45 kda. The molecule consists of a polypeptide with up to two phosphate groups per mole and a side chain of mannose and glucosamine residues. Ovalbumin is the main protein found in egg white, making up 60-65% of the total protein. Medicinal characteristics: In cases where poisoning by heavy metals (such as Iron) is suspected, ovalbumin may be administered. Ovalbumin chelates to heavy metals and traps the metal ions within the sulfhydryl bonds of the protein. Chelating prevents the absorption of the metals into the gastrointestinal tract and prevents poisoning.

37 Contractile proteins are proteins which participate in contractile processes. They include muscle proteins as well as those found in other cells and tissues. These proteins participate in localised contractile events in the cytoplasm, in motile activity, and in cell aggregation phenomena.

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39 The cytoplasm of cells is a colloidal network of contractile proteins. Actin filaments are the major components of this network. Other contractile proteins interact with these filaments to create structural rigidity and movement. The structure and function of contractile proteins is striated muscles is well characterized and thus provides a good example for extrapolitation to an analysis of contractileprotein structure and function of nonmuscle cells. However, the interaction of contractile proteins of various cells may be unique.

40 The study of contractile proteins in cells other than muscle has distinct difficulties: (a) The proteins are present in much lower concentration than in muscle, and only a few cell types are obtainable for study in quantities comparable to muscle. (b) Proteolysis and other detriments may be more severe in nonmuscle cells. (c) The organization of contractile proteins is difficult to define in nonmuscle cells. (d) The effort is diffuse; investigations examine a wide variety of different, or less commonly the same, cells.

41 The most prominent example of a motor protein is the muscle protein myosin which "motors" the contraction of muscle fibers in animals. Myosin is the protein responsible for generating muscle contraction. Many molecules of myosin generate enough force to contract muscle tissue. Myosins are also vital in the process of cell division.

42 Structure and Function: Most myosin molecules are composed of a head, neck, and tail domain. Domains: The head domain binds the filamentous actin, and uses ATP hydrolysis to generate force and to "walk" along the filament towards the (+) end (with the exception of one family member, myosin VI, which moves towards the (-) end). The neck domain acts as a linker and as a lever arm for transducing force generated by the catalytic motor domain. The neck domain can also serve as a binding site for myosin light chains which are distinct proteins that form part of a macromolecular complex and generally have regulatory functions.

43 The tail domain generally mediates interaction with cargo molecules and/or other myosin subunits. In some cases, the tail domain may play a role in regulating motor activity. The thick myosin bands are not single myosin proteins but are made of multiple myosin molecules. Each myosin molecule is composed of two parts: the globular "head" and the elongated "tail". They are arranged to form the thick bands

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