Proteins Lecture Notes PDF

1 2 1  Lecture Learning Outcome/Outline Students should be able to identify/describe:  Function & types of proteins, General features of proteins & Structures of amino acids  Levels of protein structures...

1 2 1  Lecture Learning Outcome/Outline Students should be able to identify/describe:  Function & types of proteins, General features of proteins & Structures of amino acids  Levels of protein structures (primary, secondary, tertiary & quartenary  Amino acid metabolism: -Transamination & oxidative deamination - UREA cycle  Nitrogen containing compounds 3  Enzymes ◦ accelerate chemical reactions  Transfer proteins ◦ Hemoglobins  Nutrient and storage proteins ◦ Ovalbumin, the major protein of egg white ◦ casein, major protein in milk  Contractile or motile proteins ◦ Tubulin  Structural proteins ◦ collagen, leather/skin, ligament, hair, silk fiber, fibroin (spider webs)  Defense proteins ◦ thrombin and fibrinogen, blood clotting proteins. Snake venoms, bacterial toxins etc.  Regulatory proteins ◦ Hormones, insulin regulates sugar metabolism; G proteins, which mediate cellular response to many hormonal signals. 4 2   Proteins are large  Proteins have characteristic amino acid compositions  Some proteins contain chemical groups other than amino acids :– conjugated proteins ◦ lipoprotein, glycoprotein, metalloprotein ◦ prosthetic group 5 Proteins lose structure and function upon denaturation. Denaturation agents:  heat  extremes of pH  organic solvents such as alcohol or acetone, by solute (urea). 6 3  http://study.com/academy/lesson/dipeptide-definition-formation-structure.html 7 The nonionic form does not exist in aqueous conditions 8 4 9 http://www.easybiologyclass.com/mcq-sample-questions-biochemistry-amino- acids-csir-icmr-jrf-net-life-science-examination/ 10 5 11 OXIDIZED Disulphide bonds in insulin molecule 12 6 13 Ornithine and citrulline are intermediates in biosynthesis of urea 14 7  glycine glutamate histidine 15 16 8  1° Primary – sequence of amino acids 2° Secondary - local structures - H-bonds 3° Tertiary - overall 3-dimensional shape super-secondary structure independent globular folding unit 4° Quaternary - subunit organization https://www.youtube.com/watch?v=1peFJ_-N7V8 17 Reading frame-sequence of codons from start to stop specifying order of amino acid Primary structure is the amino acid sequence Numbering of amino acid: From N terminal 18 9  Protein Secondary Structure – alpha helices beta pleated sheets 19 Tertiary structure - fold of a given chain Quaternary structure - protein functional unit 20 10  Fibrous – Globular – Membrane – extended, compact, part lipid soluble insoluble soluble 21 Keratin Collagen Fibrous proteins Silk Elastin http://netdugout.com/2016/10/global-silk-sales-market-2016-industry-revenue- http://shilpsnutrilife.blogspot.my/2013/05/keratin-protein-in-your-hair-nail- growth-supply-consumption-forecast-2022/ and.html 22 11   Repeating amino acid sequence  Unusual amino acid composition  One kind of secondary structure  Often mechanically strong  Usually insoluble  Usually play a structural role 23 24 12   Found in hair, fingernails, skin, claws, horns and beaks  Sequence consists of 311-314 R-hand residue -helical rod segments  Unusual 7-residue repeats: (a-b-c- d-e-f-g)n, where ‘a’ and ‘d’ are L-hand nonpolar.  Helical rods  Held together by ‘hydrophobic interactions’  Cross-linked by disulfides (Cys- Cys)  nails have more Cysteine than hair. Therefore more rigid R-hand 25 1 strand 2 strands 4 strands 26 13  Gly Ala Gly Ala Fibroin – -pleated sheet protein – Alternating sequence: Gly-(Ala/Ser)-Gly-(Ala/Ser).... – Gly on one side – Ala or Ser on the other – Gly on one sheet meshes with Gly on an adjacent sheet (same for Ala/Ser) 27  Principal component of connective tissue (tendons, cartilage, bones, teeth) – basic molecule is tropocollagen  three intertwined helical chains (1000 residues each) – more extended than -helices – each is left handed  stabilised by H-bonds  Repeating triplet: -(Gly-Pro-(Pro/HyP))-  molecules covalently cross-linked via modified Lys residues 28 14   Unusual amino acids found: H-bonds ◦ 4-hydroxyproline prolyl hydroxylase ◦ 3-hydroxyproline Fe2+ & Vit.C ◦ 5-hydroxylysine – Lysyl oxidase (Copper) ◦ Pro and HyP together make 30% of residues Lysine residues are also post- translationally modified to aldehyde derivatives by a copper-dependent enzyme called lysyl oxidase. Two lysines can then form a very stable covalent intramolecular crosslink between two strands in tropocollagen via an aldol condensation and dehydration. Cross links 29 URAK1  Every third residue faces the crowded centre of the helix - only Gly fits here  Pro and HyP suit the constraints  Interchain H-bonds involving HyP stabilise helix  Fibrils are further strengthened by intrachain lysine-lysine & other cross-links The tight packing in the centre of the triple helix requires Gy to be present at every 3rd position. 30 15 Slide 30 URAK1 UMAH RANI A/P KUPPUSAMY, 29/11/2020  Tissue Arrangements Tendon Parallel bundles Skin Sheets of fibrils layered at many angles Cartilage No distinct arrangement Cornea Planar sheets stacked crosswise – minimizes light scatter Mammals have 30 genetically distinct polypeptide chains comprising 16 collagen variants that occur in different tissues 31 Skurvy: vitamic C deficiency inactivates prolyl hydroxylase - hydroxyproline (HyP) cannot be synthesized and the collagen is less stable – denatures to gelatin. Historically, consumption of citrus fruits (rich in Vitamin C) could prevent scurvy among sailors who often suffered from scurvy. http://www.blatner.com/adam/consctransf/historyofmedicine/5-deficiencydiseases/5-deficdis.html 32 16  Diseases of collagen (continued) Copper deficiency leads to inactivity of the cross-linking enzyme, lysyl oxidase resulting in unstable collagen mutations in collagen can result in rare heritable disorders such as osteogenesis imperfecta (brittle bone disease). https://www.dovepress.com/col1a2-gene-analysis-in-a-czech-osteogenesis-imperfecta-patient-a-cand-peer-reviewed-fulltext-article-AGG 33  Rubber-like elastic properties  Principal component of elastic connective tissue in lungs, large blood vessels & elastic ligaments  Distinct amino acid composition  1/3 Gly, 1/3 Ala+Val  lots of Pro – little HyP  Forms 3D network of fibres with no regular structure  Extensive cross-linking of Lys 34 17  This implies Anfinsen, 1957 that primary Demonstrated that proteins can fold spontaneously structure into their native conformations under physiological determines conditions. folding or 3-D stucture. Physical properties of protein that influence stability & therefore, determine its fold: ◦ Rigidity of backbone ◦ Amino acid interaction with water ◦ Hydropathy index for side chains ◦ Interactions among amino acids  Electrostatic interactions  Hydrogen, disulphide bonds  Volume constraints http://slideplayer.com/slide/8616932/ 35 Chaperone proteins Assist folding Prevent degradation Prevent aggregation & prion formation Assist translocation of some proteins What is a prion disease? http://biosocialmethods.isr.umich.edu/epigenetics-tutorial/epigenetics-tutorial-gene-expression-from-dna-to-protein/ 36 18 37 -Not required in diet -Required in diet -Can be formed from a-keto -Humans incapable of acids by transamination forming requisite carbon skeleton & subsequent reactions Arginine* Lysine Alanine Glycine Histidine* Methionine Asparagine Proline Isoleucine Threonine Aspartate Serine Leucine Phenylalanine Glutamate Cysteine(from Met*) Valine Tryptophan Glutamine Tyrosine (from Phe*) * Essential amino acids * Essential in children, not in adults 38 19   Metabolized to -ketoglutarate, pyruvate, oxaloacetate, fumarate, or succinyl CoA Aspartate Methionine Alanine Asparagine Valine Serine Arginine Glutamine Cysteine Phenylalanine Glutamate Glycine Tyrosine Proline Threonine Isoleucine Histidine Tryptophan 39  Metabolized to acetyl CoA or acetoacetate Isoleucine Lysine Leucine Phenylalanine Threonine Tyrosine Tryptophan 40 20  Amino Acid1 +-Keto Acid2 Amino Acid2 +-Keto Acid1 NH3+ O - O 2 CCH 2 CH 2 CHCO 2 - + R-CCO 2 - Glutamate Pyridoxal phosphate (PLP)- Dependent Aminotransferase O NH2 - O 2 CCH 2 CH 2 CCO 2 - + R-CHCO 2 - -Ketoglutarate 41 Glutamate Glutamate-Pyruvate Aminotransferase -Ketoglutarate + + Pyruvate Alanine Glutamate-Oxaloacetate Glutamate Aminotransferase -Ketoglutarate + + Oxaloacetate Aspartate Blood levels of these aminotransferases, also called transaminases, are important indicators of liver or cardiac disease 42 21   The amino group of an amino acid is transferred to the enzyme (aminotransferase) producing keto acid and the aminated enzyme.  The amino group (on the enzyme) is then transferred to the keto acceptor (eg. α-ketoglutarate). This produces the new amino acid (glutamate) and regenerates the enzyme.  The enzyme requires a co-enzyme (pyridoxal-5’-phosphate (PLP); derived from pyridoxine (Vit.B6)  Muscle aminotransferases accept pyruvate as their α-keto acid substrate to form alanine.  Alanine can be transported to the liver where it undergoes transamination to produce pyruvate (can be used in gluconeogenesis). The glucose produced can reach the muscle via the blood circulation.:- Glucose-alanine cycle Recall ‘Cori Cycle’ 43 The amino group of most amino acids are consequently converted to (funneled) the formation of glutamate or aspartate.  Glutamate and aspartate are interconvertible Glutamate Glutamate-aspartate α-ketoglutarate + aminotransferase + Oxaloacetate aspartate Oxidative deamination Oxidative deamination of glutamate regenerates α-ketoglutarate + NH3 Catalyzed by glutamate dehydrogenase (The only known enzyme that utilizes either NAD+ or NADP+ as its redox coenzyme) This enzyme is inhibited by GTP and activated by ADP. The resulting NH3 formed is toxic! 44 22  Note:  Glutamate dehydrogenase is the major enzyme responsible for ammonium production.  Ammonium is toxic (Normal: 15 - 40M , max 70M)  Liver: Principle site  Hyperammonium >500M plasma [NH4+] = TOXIC  related to inborn errors of metabolism (genetic defects) as well as induced (liver failure) 45 Urea Cycle The urea cycle was the first metabolic process to be described as a cycle bySir Hans Krebs who also described the TCA cycle. Role of Urea cycle: rid the body of toxic NH4+ Liver- major site of urea synthesis, as it is the major source of arginase, and is the only tissue with the complete set of all 5 enzymes required. 4 5 3 2 1 http://www.diapedia.org/metabolism-insulin-and-other-hormones/5105758814/amino-acid-metabolism 46 23  Proteolysis & amino acid catabolism (various tissues) Gut bacteria activity http://www.gohomeworkhelp.com/what-is-the-function-of-the-urea-cycle.html 47 LINK BETWEEN UREA CYCLE TCA CYCLE https://www.studyblue.com/notes/note/n/nitrogen-metabolism/deck/7035145 48 24  Nitrogen containing compounds 49 Glutamate NH3+ NH3+ decarboxylase - - O 2 CCH 2 CH 2 CHCO 2 - O 2 CCH 2 CH 2 CH 2 Glutamate Gamma-aminobutyrate CO2 (GABA) http://www.slideshare.net/dryuktisharma/action-potentials-animal-systems GABA is an important inhibitory neurotransmitter that triggers opening of chloride channels in the postsynaptic neuron, which hyperpolarizes its membrane. Drugs (e.g., benzodiazepines) that enhance the effects of GABA are useful in treating epilepsy 50 25  NH 2 NH 3 + + Arginine - H 2 N=C-HN CH 2 CH 2 CH 2 CHCO 2 Nitric oxide synthase (NOS) NH + 3 - NH 2 CONH CH 2 CH 2 CH 2 CHCO 2 + NO Citrulline 51 3-phophoglycerate CO 2 - CO 2 - dehydrogenase Glycolysis Glucose H C OH C=O NAD+ NADH + CH2 OPO3-2 H+ CH2 OPO3-2 3 steps 3-Phospho- 3-Phospho- glycerate hydroxypyruvate Pyruvate inhibit Inhibits Glutamate Transaminase -Ketoglutarate CO 2 - CO 2 - 3-phosphoserine phosphatase H C NH3 + H C NH3 + CH2 OH CH2 OPO3-2 Serine (Ser) 3-Phosphoserine 52 26  H Dihydrofolate H2N N N reductase CO 2 - Folate N N CH2 NHR OH H C NH3 + H Serine Tetrahydrofolate CH2 OH (FH4) Serine hydroxymethyl H transferase (PLP-dep.) N Key intermediate CO 2 - in biosynthesis of N CH2 Glycine H C NH3 + purines and N formation of H2 C H thymine Important in N5, N10-Methylene FH4 biosynthesis of heme, porphyrins, and purines 53 Glycine is required for Heme synthesis Glycine + Succinyl CoA δ-aminolevulinic acid synthase Protoporphyrin IX CO2 Fe2+ HEME Aminolevulinic acid (ALA) Protoporphyrinogen oxidase Mitochondria Porphobilinogen Protoporphyrinogen IX synthase Cytosol Porphobilinogen (PBG) 2CO2 Coproporphyrinogen Uroporphyrinogen oxidase 4NH3 III cosynthase Uroporphyrinogen Uroporphyrinogen III decarboxylase Coproporphyrinogen III 4 CO2 54 27   Deficiency of enzyme(s) in the heme synthesis pathway eg.  i) ferrocheletase  ii) uroporphyrinogen III cosynthase  Leads to build-up of porphyrin or other intermediates which is/are then secreted in the urine (red coloured urine)  Can affect the nervous system (neurological disorders) or skin http://ashlandscience.shoutwiki.com/wiki/Cutaneous_Porphyria_(Vampires_Disease) 55 Methionine Synthase NH3 + NH3+ (Vit. B12-dep.) + 5-Methyl FH4 - (Derived from - HSCH 2CH 2CHCO 2 CH 3SCH 2CH 2CHCO 2 + FH4 folic acid ) L-Homocysteine Methionine CO 2 - (Essential) Cystathionine H C NH3+ -synthase CH2 OH (PLP-dep.) Serine Cystathionine NH3+ OH NH3 + lyase SCH 2CH 2CHCO 2 - CH 3CHCH 2CO 2 - + HSCH 2CHCO 2 - CH2CHCO2- Cystathionine  -Hydroxy- Cysteine NH3+ butyrate (Non-essential) 56 28  Homocysteinuria Rare; deficiency of cystathionine -synthase Dislocated optical lenses Mental retardation Osteoporosis Cardiovascular disease death High blood levels of homocysteine is associated with cardiovascular disease May be related to dietary folate deficiency 57 NH2 N N S-Adenosyl methionine N N NH3 + synthase + - -O 2 CCHCH2 CH2 -S-H2 C CH 3SCH 2CH 2CHCO 2 O NH3 + CH3 Methionine ATP SAM OH OH NH2 Decarboxylase N S-Adenosyl N Methionine NH2 (SAM) N N N N + H3NCH2CH2CH 2-S-H2C O CO2 N R-H N CH3 Methyl- - transferases O2 CCHCH2CH2-S-H2 C O OH OH NH3 + R-CH3 Decarboxylated SAM (S-Adenosylmethioninamine) OH OH S-Adenosyl homocysteine 58 29 URAK2 Arginine-glycine transamidinase NH2 NH2 NH3 + (Kidney) + + H2 N=C-HNCH 2 CH 2 CH 2 CHCO 2 - H2 N=C-HNCH 2 CO 2 - Glycine Ornithine Arginine Guanidoacetate SAM + ATP H Guanidoacetate N O Methyltransferase S-Adenosyl- (Liver) homocysteine Creatinine HN + ADP Non-enzymatic (Urine) N (Muscle) CH3 NHPO3-2 + Creatine kinase H2 N=C-NCH 2 CO 2 - NH2 (Muscle) CH3 + Creatine H2 N=C-NCH 2 CO 2 - ADP + Pi Phosphocreatine CH3 ATP 59 Creatinine: Urinary excretion generally constant; proportional to muscle mass Creatinine Clearance Test: Compares the level of creatinine in urine (24 hrs.) with the creatinine level in the blood Used to assess kidney function Important determinant in dosing of several drugs in patients with impaired renal function 60 30 Slide 59 URAK2 UMAH RANI A/P KUPPUSAMY, 29/11/2020  + NH3 H N CH2 CHCO2- Histidine H decarboxylase N CH2CH2NH2 N Histidine CO2 N Histamine Histamine: Synthesized in and released by mast cells H1 receptors Mediate allergic response vasodilation, bronchoconstriction H1 blockers: Diphenhydramine (Benadryl) Loratidine (Claritin) H2 receptors Stimulates secretion of gastric acid () H2 blockers: Cimetidine (Tagamet); ranitidine (Zantac) 61 Phenylalanine and Tyrosine (A class of monooxygenase) https://es.slideshare.net/ashokktt/metabolism-of-phenylalanine-and-tyrosine/11 62 31   Deficiency of Phenylalanine hydroxylase  Occurs in 1:16,000 live births in U.S.  Seizures, mental retardation, brain damage  Treatment: limit phenylalanine intake  Screening of all newborns mandated in all states Tyr Phe Transamination Phenylpyruvate O (urine) CH2 CCO2- 63 HO HO Transamination O NH3 + CH2 CHCO2 - CH2CCO2- Tyrosine p-Hydroxyphenyl- pyruvate Deficient in O2 alkaptonuria p-Hydroxyphenyl- pyruvate OH Homogentisate dioxygenase dioxygenase (ascorbate-dep.) Cleavage of aromatic ring CO2 O2 CH2CO2- Fumarate + acetoacetate OH Homogentisate 64 32   Deficiency of homogentisate dioxygenase  Urine turns dark on standing (oxidation of homogentisic acid)  Asymptomatic in childhood  Tendency toward arthritis in adulthood http://slideplayer.com/slide/5273233/ 65 Catechol HO HO Tyr hydroxylase NH3 + NH3 + CH2 CHCO2 - O2 HO CH2 CHCO2- Tyrosine Dihydroxyphenylalanine (DOPA) HO DOPA Epinephrine (Adrenaline) decarboxylase CO2 HO CHCH2 NHCH3 HO OH Methyl Dopamine transferase hydroxylase S-Adenosyl- HO HO CH2CH2NH2 homocysteine SAM Dopamine HO CHCH2NH2 DOPA, dopamine, norepinephrine, Norepinephrine OH and epinephrine are all neurotransmitters 66 33  HO MAO HO (in mitochondria) HO CHCH2NHR' CHCHO HO R R R R’ Aldehyde dehydrogenase OH H Norepi HO OH CH3 Epi H H Dopamine CHCO2H HO R MAO inhibitors (e.g., tranylcypromine) are useful Urinary in the treatment of depression metabolite R=OH Vanillylmandelic acid (VMA) R=H Homovanillic acid (HVA) 67 HO HO COMT HO CHCH2NHR' CH3O CHCH2NHR' SAM S-Adenosyl- R homocysteine R Active Inactive catecholamine metabolite COMT found in cytoplasm Terminates activity of catecholamines Catecholamine excretion products result from combined actions of MAO and COMT Inhibitors of COMT (e.g., tolcapone) useful in Parkinson’s disease 68 34  HO HO NH3 + Tyrosinase NH3 + CH2 CHCO2 - HO CH2 CHCO2- Tyrosine DOPA Tyrosinase O Melanin Highly colored polymeric (Black polymer) O CH2 CHCO2- intermediates + NH3 Melanin formed in skin (melanocytes), eyes, and hair Dopaquinone In skin, protects against sunlight Albinism: genetic deficiency of tyrosinase 69 + Indole ring NH3 + NH3 CH2 CHCO2 - CH2 CHCO2 - CH2 CH2 NH2 Trp hydroxylase HO HO Decarboxylase N H O2 N N H H Tryptophan 5-Hydroxy- CO2 5-Hydroxy- (Trp) tryptophan tryptamine (5-HT); Serotonin 70 35  Serotonin formed in: Brain (neurotransmitter; regulation of sleep, mood, appetite) Platelets (platelet aggregation) Smooth muscle (contraction) Gastrointestinal tract (enterochromaffin cells - major storage site) Drugs affecting serotonin actions used to treat: Depression Serotonin-selective reuptake inhibitors (SSRI) Migraine Schizophrenia Obsessive-compulsive disorders Chemotherapy-induced emesis (nausea / vomiting) Some hallucinogens (e.g., LSD) act as serotonin agonists 71 CH2 CH2 NH2 CH2CHO HO MAO HO N N H H Serotonin Dehydrogenase CH2CO2H HO Carcinoid tumors: Malignant GI tumor type N Excretion of large amounts of H 5-HIAA 5-Hydroxyindole acetic acid (5-HIAA) (Urine) 72 36  Serotonin- N acetyl transferase Hydroxyindol-o- methyltransferase Melatonin: Formed principally in pineal gland Brain produces in response to darkness It helps with circardian rhythms 73 + NH3 CH2CHCO2 - CO2 H Several steps N N H Tryptophan Nicotinic acid (Niacin) Nicotinamide adenine dinucleotide (NAD) 74 37  Task 16 Urea cycle Task 17 Prepare a table to summarise amino acid, its role/relevance in our body and how it can be metabolised. 75 76 38 Here’s an elaboration of the document’s content in the correct sequence: 1. Protein Functions and Types Proteins ser ve various roles in the body: Enzymes: Accelerate chemical reactions (e.g., digestive enzymes). Transfer Proteins: Carry molecules like oxygen (e.g., hemoglobin). Nutrient and Storage Proteins: Provide energy or nutrient reser ves (e.g., casein in milk). Contractile Proteins: Enable movement (e.g., tubulin). Structural Proteins: Provide support (e.g., collagen in tendons). Defense Proteins: Protect against threats (e.g., antibodies, blood clotting factors). Regulatory Proteins: Control processes like sugar metabolism (e.g., insulin). 2. General Features of Proteins Composition: Made of amino acids; some include prosthetic groups (lipoproteins, glycoproteins). Denaturation: Loss of function due to heat, pH extremes, or solvents like alcohol. 3. Protein Structure Levels Primary Structure: Linear sequence of amino acids. Secondary Structure: Local folding patterns like alpha-helices and beta-pleated sheets (held by hydrogen bonds). Tertiary Structure: The overall 3D shape of a single polypeptide. Quaternary Structure: Arrangement of multiple subunits in a functional protein (e.g., hemoglobin). 4. Amino Acid Metabolism Transamination: Transfer of amino groups bet ween amino acids and keto acids (e.g., alanine to pyruvate). Oxidative Deamination: Removes amino groups, releasing ammonia (toxic if not processed). Urea Cycle: Converts toxic ammonia into urea for safe excretion. 5. Structural Proteins Fibrous Proteins: Provide mechanical support (e.g., collagen, keratin). Collagen: Found in connective tissues, stabilized by cross-links and hydrogen bonds. Keratin: Found in hair, nails; rich in cysteine, forming disul de bonds. Fibroin: Main component of silk, structured in beta-pleated sheets. Globular Proteins: Compact and soluble, often enzymes or hormones. Membrane Proteins: Embedded in lipid membranes, playing roles in transport or signaling. 6. Metabolism of Amino Acids Essential Amino Acids: Required from diet (e.g., lysine, valine). Non-Essential Amino Acids: Synthesized by the body (e.g., alanine, glutamate). Degradation Pathways: Amino acids are broken down into intermediates for energy (e.g., acetyl-CoA, pyruvate). 7. Nitrogen-Containing Compounds Neurotransmitters: Derived from amino acids. GABA: From glutamate, inhibits nervous system activity. Serotonin: From tryptophan, regulates mood and sleep. Dopamine: Precursor to norepinephrine and epinephrine. Heme Synthesis: Glycine is essential for forming heme, a component of hemoglobin. 8. Diseases Related to Protein and Amino Acid Metabolism Scurvy: Vitamin C de ciency affects collagen stability. Phenylketonuria (PKU): De ciency of phenylalanine hydroxylase leads to cognitive issues; treated by dietary restrictions. Alkaptonuria: Caused by homogentisate oxidase de ciency; results in darkened urine and joint issues. Homocystinuria: High homocysteine levels linked to cardiovascular disease. This over view summarizes the document in sequence. Let me know if you want details on any speci c topic! fi fi fi fi fi