Amino Acids and Proteins Chapter 9 PDF

Amino Acids and Proteins CHAPTER 9 Preamble PowerPoints are a general overview and are provided to help students take notes over the video lecture ONLY. PowerPoints DO NOT cover the details needed for the Unit exam Each student is responsible for READING the TEXTBOOK for details to ans...

Amino Acids and Proteins CHAPTER 9 Preamble PowerPoints are a general overview and are provided to help students take notes over the video lecture ONLY. PowerPoints DO NOT cover the details needed for the Unit exam Each student is responsible for READING the TEXTBOOK for details to answer the UNIT OBJECTIVES Unit Objectives are your study guide (not this PowerPoint) Test questions cover the details of UNIT OBJECTIVES found only in your Textbook! Introduction Proteins are complex polymers of α-amino acids that are produced by living cells in all forms of life. Each protein is composed of a maximum of 20 different amino acids in varying numbers and sequences. All proteins contain carbon, hydrogen, oxygen, and nitrogen, and some include sulfur. Protein Structure (1) Amino acids contain an amino group (NH2), carboxyl group (COOH), hydrogen, and an R group (radical or side chain) with the formula RCH (NH2 ) COOH. Protein Structure (2) The structure of amino acids is amphoteric-containing two ionizable sites, a proton-accepting group (NH2) and a proton-donating group (COOH). When both are ionized, the amino acid is called an ampholyte or dipolar ion. The isoelectric point (pI) is the pH at which the amino acid or protein has no net charge and the positive charges equal the negative charges. At a pH greater than the pI, the protein carries a negative charge and at a pH less than the pI, the protein carries a positive charge. Protein Structure (3) Peptide Bond A molecule of water is split between the carboxyl group of one amino acid and the amino group of another and a covalent bond called a peptide bond. The end of the protein that has the amino-free group is called the N-terminal end and the opposite end which has the carboxyl-free group, is called the C- terminal end.  Protein Structure (4) Primary Structure The sequence of amino acids in the polypeptide chain-the identity and specific order of the amino acids. Secondary Structure Determined by the interaction of adjacent amino acids. Three possible conformations are the α-helix, β-pleated sheets, and random coils.  Protein Structure (5) Tertiary Structure The way in which the chain folds back upon itself to form a 3-dimensional structure. These are mainly interactions of amino acids with the R-groups of more distant amino acids. Determines the chemical and physical properties of the protein. Quaternary Structure The arrangement of two or more polypeptide chains to form a protein. Only proteins with more than one polypeptide chain have this. Protein Structure (6) Denaturation Disruption of the bonds holding the secondary, tertiary, or quaternary structures together Can occur as a result of heat, changes in pH, mechanical forces, exposure to chemicals (solvents, detergents, metals), and exposure to ultraviolet light.  Protein Metabolism Digestion of dietary proteins by proteolytic enzymes Originates in the gastrointestinal tract Liver and other organs utilize the amino acid pools to synthesize the body’s proteins. In the kidneys, amino acids are filtered through the renal glomeruli. Subsequently reabsorbed by the renal tubules Protein Synthesis Most plasma proteins are synthesized in the liver and secreted into circulation by the hepatocytes. Double-stranded DNA molecule unfolds. One strand serves as a template for the messenger RNA (mRNA). Code is carried by the mRNA from nucleus to cytoplasm. Attaches to a ribosome receptor protein. Amino acid linked to transfer RNA (tRNA) that corresponds to the specific codon is carried to the ribosome and is attached to the matching codon. Next amino acid in the sequence is added. Cycle repeats itself until the protein is completed. Protein Functions Maintenance of water distribution between cells and tissue. When protein levels are decreased, the osmotic pressure is also decreased. Coagulation proteins are important in maintenance of hemostasis. Many proteins function as transport vehicles to move various ligands to where they are needed or stored. Box 9-1 Protein Functions Maintenance of colloidal osmotic pressure and water distribution Structural-support for the body, tissue, or cell Collagen Keratin-hair, nails Transport molecule; for example, Transferrin- Fe+3 Albumin-bilirubin Thyroid binding globulins Hormones Enzymes Peptide hormones, insulin Coagulation Hemoglobin Antibodies  Aminoacidopathies (1 ) Inherited disorders of amino acid metabolism Over 100 aminoacidopathies have been identified. 1. Alkaptonuria Rare inherited disease involving the homogentisic acid oxidase (H G O) Leads to a buildup of homogentisic acid (H G A) in the tissues of the body Autosomal recessive condition Ochronotic Darkening of the tissues of the body because of the excess homogentisic acid 2. Cystinuria Defect in the amino acid transport system Cystine is somewhat insoluble. Resulting in precipitation in the renal tubules, formation of urinary calculi Aminoacidopathies (2) 3. Maple Syrup Urine Disease Named for characteristic maple syrup or burnt sugar odor of the urine Caused by the absence or very low levels of the branched-chain enzyme α-ketoacid decarboxylase complex Results in the abnormal metabolism of three essential amino acids: leucine, isoleucine, and valine Treatment involves a special, very carefully controlled diet requiring careful monitoring of protein intake. 4. Phenylketonuria Inborn error of metabolism Results in the inability to metabolize the essential amino acid phenylalanine Autosomal recessive trait Pregnant women known to be carriers of the P K U gene or definitely carrying a P K U fetus should also be maintained on a phenylalanine-restricted diet from conception to birth. Elevated phenylalanine levels are toxic to developing brain tissue and negatively impact brain function. Specific Plasma Proteins (1) Proteins are categorized into two main groups: 1. Albumin 2. Globulin 1. α1 – globulins 2. α2 - globulins 3. β – globulins 4. γ – globulins Specific Plasma Proteins (2 ) Prealbumin/Transthyretin TTR binds with thyroxine and triiodothyronine (thyroid hormones) and retinol (vitamin A). Serves as a transport protein Main clinical significance of prealbumin is role as a sensitive marker of poor nutritional status such as protein-energy malnutrition (PEM). Decreased prealbumin indicates dietary intake of protein is not adequate. Results in decreased synthesis of prealbumin by the liver. Also decreased in: Acute inflammatory response (acute phase reactant, APR) Liver disease Nephrotic syndrome Other protein-losing renal diseases Specific Plasma Proteins (3) Albumin Synthesized in the liver Comprises approximately 60% of total serum protein Chief biological function is to maintain plasma colloidal osmotic pressure (COP). Another function is to transport and store a wide variety of ligands. Hypoalbuminemia Decreased albumin levels Most common cause is increased catabolism due to tissue damage and inflammation. Hyperalbuminemia Little diagnostic significance except in dehydration. Increase is usually artifactual due to a decrease in plasma volume Functions of Albumin *Maintain plasma colloidal osmotic pressure Bind and transport a wide variety of ligands Bilirubin Long chain fatty acids Therapeutic drugs (e.g., warfarin, diazepam, digoxin, phenylbutazone, salicylate, penicillin) Calcium Magnesium Hormones (e.g., thyroxine, triiodothyronine, cortisol) Serve as an endogenous source of amino acids Acid base balance Pro- and anti-coagulatory effects *Chief biological function Hypoalbuminemia (1 of 2) *Increased catabolism: tissue damage and inflammation Impaired or decreased synthesis Primary: liver disease Secondary: diminished protein intake, malnutrition, malabsorption Increased loss of protein Nephrotic syndrome Chronic glomerulonephritis Diabetes mellitus/diabetic nephropathy Extensive burns Acute viral gastroenteritis *Most common cause  Globulins (1) a1 Globulins a1 -antitrypsin a1 (AAT) is the major a1- globulin, making up approximately 90% of a1-proteins It is an acute phase reactant (A P R) with antiprotease activity resulting in the neutralization of leukocyte elastase and collagenase. A A T deficiency is one of the most common genetically lethal diseases in Caucasians (1:4000). Emphysema with onset at 45 years of age or earlier or emphysema occurring in the absence of smoking are common features of A A T deficiency. A congenital deficiency can also result in juvenile hepatic cirrhosis, where A A T is synthesized by the hepatocytes but not released. Globulins (2) a1 Globulins a1-Acid Glycoprotein (Orosomucoid) AAG is the major glycoprotein increased during inflammation, an APR. Elevated levels are found in rheumatoid arthritis, cancer, pneumonia, and other conditions resulting in an APR. Alpha-Fetoprotein (A F P) Principal fetal protein (fetal albumin-like protein) in maternal serum Used to screen for the antenatal diagnosis of neural tube defects including spina bifida and anencephaly AFP is decreased in Down’s syndrome and Trisomy 18. Can also be used as a tumor marker, with increased levels found in 80% of patients with hepatocellular cancer, 50% of germ cell tumors (gonadal), and all children with hepatoblastoma. Globulins (3) a2 Globulins Haptoglobin (Hp) An acute phase reactant that binds free hemoglobin in plasma Prevents loss of hemoglobin and its iron through the renal glomeruli. Another important role is control of local inflammatory response through a number of processes Depletion is the most sensitive indicator of intravascular hemolysis, in transfusion reactions, and certain hemolytic disorders (hemolytic anemias). Is an APR that is synthesized late and is weak reacting Is increased in conditions involving inflammation, infection, tissue necrosis, or malignancy Globulins (4) a2 Globulins Ceruloplasmin (Cp) Principal copper (Cu)-containing protein in plasma containing 95% of the total serum copper Although Cu is an essential nutrient, it is very toxic to cells in high concentrations. Primary storage site is the liver. Principal site of excretion is the biliary tract. Globulins (5) a2 Globulins Ceruloplasmin (Cp) Wilson Disease (WD) Rare autosomal recessive trait where Cp levels are reduced and the dialyzable Cu concentration is increased. Excessive accumulation of Cu in the liver, kidney, and brain can lead to: Degenerative cirrhosis Chronic active hepatitis Renal tubular acidosis Neurological damage (clumsiness, tremors) unless treated with a copper chelator, for example, penicillamine or trientine. Cu deposits in the eyes, resulting in the characteristic Kayser-Fleischer rings, pigmented rings at the outer margins of the cornea and the sclera. Cp is an acute phase reactant that increases late in the condition. Globulins (6) a2 Globulins a2-Macroglobulin (AMG) One of the largest plasma proteins In nephrotic syndrome, AMG is characteristically increased up to 10 times normal because it is retained while smaller proteins are excreted in the urine. Elevated in liver disease and estrogen [oral contraceptives or hormone replacement therapy (HRT)] and slightly increased in diabetes mellitus Globulins (7) β-Globulins Transferrin (TRF) Major component of β-globulins Principal plasma protein for transport of iron (Fe+3 – Ferric Ion) to storage sites, where it is bound to apoferritin and stored as ferritin Important in the differential diagnosis of anemias and monitoring the treatment of iron deficiency anemia when the transferrin level is increased but the percent saturation is decreased Also increased in hepatitis, pregnancy, and women on oral contraceptives or HRT A negative acute phase reactant A commonly used indicator of iron overload-screen for hemochromatosis Globulins (8) β-Globulins Hemopexin Removes heme from circulation When red blood cells are destroyed, hemopexin transports heme to the liver, where it is catabolized by the reticuloendothelial system. Increased levels are found in pregnancy and in diabetes mellitus. β-Lipoproteins Classified as low-density lipoproteins (LDLs) Transport the majority of cholesterol in the body from the liver to the tissues High levels of LDLs are a risk factor for atherosclerosis and heart disease. Globulins (9) β-Globulins β2-Microglobulin (BMG) Low-molecular-weight protein Comprises the common light chain of class I major histocompatability complex (MHC) antigens found in all nucleated cells Increased in renal failure; inflammation; and neoplasms especially those associated with B-lymphocytes Globulins (10) β-Globulins C-Reactive Protein (CRP) An acute phase reactant and a non specific indicator of bacterial or viral infection, inflammation, and tissue injury or necrosis Reacts with proteins present in many bacteria, fungi, and protozoal parasites One of the first APRs to be discovered and one of the most sensitive Levels rise dramatically following myocardial infarction, trauma, psychological or physical stress, infection, inflammation (e.g., rheumatic fever, rheumatoid arthritis), surgery, and various cancers. Has been found to be valuable in the diagnosis of bacterial infection and in differentiating between bacterial and viral infections Globulins (11) γ-Globulins Immunoglobulins or humoral antibodies Each immunoglobulin (Ig) molecule consists of two or more basic units consisting of two identical heavy (H) chains and two identical light chains (L).  Globulins (12) γ-Globulins Classes and heavy chains (idiotypes) are: IgM = μ (mu) IgG = γ (gamma) IgA = α (alpha) IgD = δ (delta) IgE = ε (epsilon) Immunoglobulin M (IgM) Largest Ig with a molecular weight of 900,000 and accounts for 5-10% of total Igs First produced during an immune response (primary response) Globulins (13) γ-Globulins Immunoglobulin G (IgG) Has a molecular weight of 150,000 Most abundant Ig in serum (70-75% of Igs) IgG antibodies are produced in response to the antigens of most bacteria and viruses. Immunoglobulin A (IgA) Has a molecular weight of 160,000 Comprises 10-15% of Ig. Secretory IgA, a dimer, is found in secretions including tears, sweat, saliva, and milk as well as gastrointestinal and bronchial secretions. Globulins (14) γ-Globulins Immunoglobulin D (IgD) Molecular weight of 184,000 Makes up 61% of serum Igs Primary function is unknown. Immunoglobulin E (IgE) Has a molecular weight of 180,000 Concentrations are very low (0.3 μg/mg). Produced in allergic reactions, urticaria, hay fever, and asthma Hyperproteinemia Associated with a positive nitrogen balance Dietary nitrogen intake is greater than the excretion or loss of nitrogen, which occurs mainly in the urine. Usually occurs as a result of hemoconcentration or dehydration Hypoproteinemia Causes a negative nitrogen balance Excretion of nitrogen exceeds intake or synthesis of protein. Most common cause is an increase in plasma water volume, or hemodilution. Relative hypoproteinemia Hyperproteinemia Hemoconcentration/dehydration Addison’s disease Inadequate intake of H2O Diabetic ketoacidosis Excessive sweating Increase in globulins Vomiting Multiple myeloma Salt-losing syndromes Waldenstrom’s Diarrhea macroglobulinemia Chronic inflammatory conditions H I V/A I D S  Hypoproteinemia Increase in plasma water volume- Blood loss after trauma hemodilution (relative hypoproteinemia) Burn patients Water intoxication Trauma Salt-retention syndromes Blood loss after trauma Massive I V infusions Burn patients Volume expanders (e.g., dextran) Trauma Increase in protein loss (kidneys, Decreased synthesis gastrointestinal tract, and skin) Liver disease Nephrotic syndrome Immunodeficiency disorders Total Protein Methodologies (1 of 5) Biuret Based on presence of peptide bonds found in all proteins. When a solution of protein (serum or plasma) is treated with cupric (Cu+2) divalent ions in a moderately alkaline medium, a violet-colored chelate, which absorbs light at 540 nm, is formed between the cupric ion and carbonyl oxygen and the amide nitrogen atoms of the peptide bond. Total Protein Methodologies (2 of 5) Biuret Biuret Reagent Sodium potassium tartrate Complexes with cupric ions to prevent their precipitation in alkaline solution Copper sulfate Major reactant providing the Cu+2 Ions Potassium iodide Antioxidant that stabilizes the cupric ions NaOH Provides the alkaline pH Total Protein Methodologies (3 of 5) Refractometry Used for a rapid, approximate measure of total serum protein concentration within the range of 3.5-10 g/dL. In plasma, the major solute protein is diluted in water and the refractive index of the solution increases in proportion to the concentration of the solute, protein. Dye-Binding Method Coomassie Blue G-250, is dissolved in an acidic solution, causing it to absorb at 465 n m. The dye, which is negatively charged, binds to the positively charged protein molecule, the absorbance undergoes a shift to 595 n m. Used to determine the protein concentration Total Protein Methodologies (5 of 5) Reference Range The serum protein in healthy, ambulatory adults in 6.0-8.3 g/dL. A physiological decrease of approximately 0.5 g/dL occurs in bed-ridden patients. A/G Ratio Globulin concentration can be calculated by subtracting the albumin from total protein. Globulin = Total Protein (g/dL) − Albumin (g/dL) The A/G ratio can then be determined by dividing the albumin concentration by the calculated globulin. Urinary Proteins Originate mostly from the blood and filtration through the renal glomeruli Proteins in the urine have been filtered by the glomeruli and have not been reabsorbed by the renal tubules. Most common method of screening for urinary protein is by urine reagent test strips. Indicator tetrabromphenol blue, at a pH of 3.0, is yellow in the absence of protein. Turns green and finally blue as the concentration of protein increases Cerebrospinal Fluid Protein CSF is a clear, colorless fluid that contains small amounts of glucose and protein. Reference range for CSF protein is 15-45 mg/dL. Increased levels of CSF proteins are found in: Various types of meningitis Encephalitis Subarachnoid hemorrhage Intracranial tumors Multiple sclerosis Guillain-Barre syndrome Brain abscesses Albumin Methodologies (1 of 2) Albumin methodologies are based on binding of albumin with anionic dyes. Bromcresol green (BCG) Bromcresol purple (BCP) The reference range for albumin is 3.4-5.0 g/dL or 34-50 g/L. Albumin Methodologies (2 of 2) Four main requirements of dye-binding methods are 1. Specific binding of the dye to albumin in the presence of serum or plasma proteins 2. High binding affinity between the dye and albumin 3. Substantial shift in the absorption wavelength of the dye in the bound form 4. Absorption maximum for the bound form at a wavelength distinct from those where bilirubin and hemoglobin, the main interfering chromogens, can interfere Protein Electrophoresis (1 of 2) Migration of charged solutes or particles in a liquid medium under the influence of an electrical field Proteins are ampholytes or zwitterions and can move toward the anode or cathode, depending on the charge. Performed on serum to avoid complication of the fibrinogen band in the β-γ region. After electrophoresis, the bands are fixed by immersing the strip in an acid medium.  Protein Electrophoresis (2 of 2) A normal serum protein electrophoresis (SPE) has five bands. The % distribution × total protein = g/dL. Relative % g/dL Albumin (anode-fastest) 53-65 3.5-5.0 Alpha sub 1 at globulin 2-5 0.1-0.3 Alpha sub 2 at globulin 7-13 0.6-1.0 β-globulin 8-14 0.7-1.1 γ-globulin (cathode-slowest) 12-20 0.8-1.6  Common Abnormal Protein Electrophoresis Patterns (1 of 3) Fusion of the β-γ Bands, or Bridging Result of fast-moving γ-globulins that prevent resolution of β- and γ-globulins Cirrhosis is the most common cause of β-γ bridging. Can also be found in chronic infections and autoimmune or collagen disorders Nephrotic syndrome Characterized by a decrease in albumin and γ-globulin bands in conjunction with an increase in a2 – globulins that suggests selective proteinuria  Common Abnormal Protein Electrophoresis Patterns (2 of 3) Fusion of the β-γ Bands, or Bridging Acute Phase Reaction Acute phase reactants that increase during APR are positive acute phase proteins (AAPs): a1-proteins (a1-antitrypsin, a1-acid glycoprotein) a2-proteins (haptoglobin, ceruloplasmin, a2–macroglobulin) β-proteins (fibrinogen, C-reactive protein) Negative AAP s are albumin, transferrin, and transthyretin (prealbumin). Common Abnormal Protein Electrophoresis Patterns (3 of 3) Fusion of the β-γ Bands, or Bridging Acute Phase Reaction Acute phase reactants are increased in: Infection Tumor growth or malignancy Rheumatoid arthritis Hepatitis Surgery Trauma Burns Myocardial infarction  Abnormal Protein Electrophoresis Patterns Multiple myeloma Cancer of the plasma cells Plasma cells produce immunoglobulins that normally protect us from infection but in multiple myeloma these are nonfunctional and are called paraproteins. Increase in the γ globulin Postamble READ the TEXTBOOK for the details to answer the UNIT OBJECTIVES. USE THE UNIT OBJECTIVES AS A STUDY GUIDE All test questions come from detailed material found in the TEXTBOOK (Not this PowerPoint) and relate back to the Unit Objectives