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Lecture (14) Peptides Proteins  Peptide bond (amide bond)  It results from a condensation reaction (dehydration) between COOH of first amino acid and NH3+ of next amino acid.  Features of the peptide bond : 1) Zigzag. ‫متعرج‬ 2) It has resonance structure (doubl...

Lecture (14) Peptides Proteins  Peptide bond (amide bond)  It results from a condensation reaction (dehydration) between COOH of first amino acid and NH3+ of next amino acid.  Features of the peptide bond : 1) Zigzag. ‫متعرج‬ 2) It has resonance structure (double bond keeps shifting position between CO and CN). 3) Partial double bond (intermediate between single and double). 4) Charged (δ+ on N and δ− on O). 5) Planar (flat) and rigid (stronger than single) → un-rotatable (fixed): - Rotation only occurs around the α-carbon within the same amino acid → moving R-groups up and down and affecting protein structure.  Note: number of peptide bonds = number of amino acid residues -1 6) Hydrogen bonding in the backbone between amine and carbonyl(except for proline): - NH with δ+ acts as hydrogen bond donor. - O with δ− in carbonyl acts as hydrogen bond acceptor. - This hydrogen bonding is responsible for stability of protein secondary structure.  Directionality of peptide: A. First amino acid always has free NH3 (called N-terminus). B. Last amino acid always has free COOH (called C-terminus). - We always read amino acids from N toward C terminus. - Addition of new amino acids occur on C-terminus during translation ‫على الذيل‬.  Backbone of peptide ‫ العامود الفقاري‬: - It consists of repeated α-amide N, α-C, and α carbonyl C atom (N-C-C - N-C-C …..). - Backbone carries the R group (but it is not involved in formation of the backbone). - R-groups mainly occur in Trans orientation (except proline which occurs in 50/ 50).  Proline as an exception 1) Proline backbone can’t act as a hydrogen bond donor: - This is because the N is a secondary amine → it is bonded to the α-carbon and the R- group → doesn’t have H → can’t act as hydrogen bond donor. - Carbonyl can still act as hydrogen bond acceptor. 2) 90% of amino acids occur in Trans orientation to decrease stearic hindrance or repulsion which is greater in the cis direction: - 50% of proline occurs in cis, and 50% in trans (no preference). - In proline, both cis and trans conformations have equal energies. - This is because proline has rigid ring and will cause stearic hindrance or repulsion in both directions ‫عاملة أزمة على الوجهين فمش فارقة معاه‬. - Proline is thus found in the cis configuration more frequently than other amino acids.  Examples of functional and exceptional peptides Carnosine (-alanyl-L-histidine) :  A dipeptide of -alanine and histidine.  The amino group is bonded to the  -carbon of alanine.  It is highly concentrated in muscle and brain tissues, with following functions: 1) Contraction of muscle. 2) Protection of cells from ROS (Radical Oxygen Species) : - ROS are free radicals that contain a single electron. - They may attack and damage all components of the cell (oxidizing agents) → cause many diseases in humans. ‫يحاولوا تعويض عقدة نقص االكترونات اللي عندهم‬ Glutathione (-glutamyl-L-cysteinylglycine):  It is a tripeptide made of glutamic acid, cysteine and glycine.  Glutamic acid uses its γ - COOH to form the peptide bond with cysteine.  The functional group is the SH of cysteine : - It scavenges ‫ تنظف‬oxidizing agents or ROS by reacting with them to protect cells. - Two molecules of the reduced glutathione (GSH) becomes oxidized → a disulfide bond (S-S) is formed between SH groups of the two cysteine residues → oxidized form of glutathione is formed (GS-SG). ‫المركب يضحي بنفسو بإعطاء الكترونات للمركبات المؤذية‬ - The reduced form (GSH) is later reformed (recycled) by special enzymes to be used again against ROS. 2 GSH GS- SG Enkephalins :  Two pentapeptides found in the brain and function as analgesics (pain relievers or killers).  The 2 pentapeptides differ only in their C-terminal amino acids: 1) Met-enkephalin: Tyr-Gly-Gly-Phe-Met 2) Leu-enkephalin: Tyr-Gly-Gly-Phe-Leu   Aromatic R-groups of tyrosine and phenylalanine play a role in their activities as analgesics.  Synthetic opiates ‫ مخدرات صناعية تعطى لتسكين األلم‬like morphine have similar 3-D structure to natural enkephalins and may bind to same receptors. Oxytocin and vasopressin (ADH):  Both are nonapeptide hormones (9 amino acid residues). They differ only at 2 residues (3rd and 8th positions).  Both have similar special features : 1) Cyclic structure due to S-S link between Cys 1 and Cys 6. 2) Both have amide group at the C-terminus (which increases stability). A. Oxytocin hormone regulates labor contraction (contraction of uterine muscle during delivery ‫)يحفز انقباض الرحم عند الوالدة‬. B. Vasopressin (ADH ‫ )الهرمون مضاد التبول‬regulates : - Contraction of smooth muscle in blood vessels → increases blood pressure. - Prevents urination → increases water retention ‫ يزيد احتباس الماء بمنع التبول‬.  Practice  Identify the 9 amino acids in the primary structure of vasopressin (ADH)? - Cysteinyl -L- Tyrosyl -……………………….L-glycine.  Please notice that the start has amine (NH2), while the end has amide CO – NH2. Refer to the figure. Which of the following is true about this peptide? a- This peptide is a pentapeptide. b- Bond Z is stronger than bond W. c- Ala is on the N-terminus and Lys is on the C- terminus. d- Alpha carbon contributes to bond Z. e- This peptide has a total positive charge at the physiological pH. Answer: D Aspartame ‫محلي صناعي‬  It is a synthetic dipeptide of L-Aspartyl-L-phenylalanine.  It is a methyl ester : C-terminus COOH is esterified to methanol to increase stability.  Aspartame is 200 times sweeter than natural sugar: - If a D-amino acid is substituted for either amino acid or for both of them, the resulting derivative is bitter rather than sweet ‫يصبح مر بدال من حلو‬.  Aspartame is used in many food products: soft drinks, candies…..  In June 2023 , WHO has declared aspartame as possible carcinogen. Phenylketonuria (PKU)  PKU is a hereditary “inborn error of metabolism” ‫خلل وراثي منذ الوالدة‬  It is caused by a defect in Phenylalanine Hydroxylase (PAH) which is the enzyme responsible for converting Phe Tyr : A. Synthesis of tyrosine derivatives may decrease, including catecholamines, melanin…. B. There is accumulation of phenylpyruvate in brains which is toxic and causes damage → causes mental retardation in children.  Management of PKU : 1) Tyrosine becomes essential for PKU patients. 2) Phenylalanine-free diet : source of phenylalanine (such as aspartame) must be limited : - Those patients should decrease intake of proteins. 3) PKU patients can be given Alitame sweetener which is a substitute for aspartame. Alitame contains alanine rather than phenylalanine : - Alitame consists of aspartic acid and alanine.  PKU is a common metabolic disorder. Newborns are commonly screened for this disease in many countries, including Jordan. Overview of proteins  Proteins have different structures (some have repeating structures, while others don’t).  Theoretically, a protein may have gazillion ‫عدد النهائي‬ possibilities of structures, but usually one is active : - These active structure is known as native conformation. - Native conformation ‫الشكل األصلي‬: 3-D structure of a properly folded and functional protein. - Protein will choose the 1 native conformation that allows maximum stability with least amount of energy. - So, when the cell makes many copies of same protein, they all take same native conformation.  Remember that peptides usually don’t have defined structure and function, but there are exceptions like glutathione, ADH, oxytocin…..  Recently, scientists were able to predict protein structure with the help of AI with high success : - This is very important for many sciences, because it helps us understand function, understand diseases, and develop drugs that target proteins based on structure.  Levels of protein structure 1) Primary structure: sequence or order of amino acid residues from N-terminus to C-terminus. ‫مجرد ذكر لألحماض األمينية بالتوالي‬ 2) Secondary structure: the localized organization of parts of a polypeptide (for example, the distinct structure formed by 10 amino acids in small part of whole protein). 3) Tertiary structure: 3-D structure and arrangement of all the amino acids residues of a single polypeptide chain: - It is how all secondary structures are organized together in 3D (whole single polypeptide). 4) Quaternary structure: describes the number and relative positions of the polypeptide subunits in a multimeric protein: - Some proteins are made of multiple polypeptides crosslinked (connected) with each other. These are known as multimeric proteins. - It is how multiple polypeptides are organized together in 3D to form 1 protein.  Primary structure  The order in which the amino acids are covalently linked together by peptide bonds: - Example: Leu—Gly—Thr—Val—Arg—Asp—His  Tissue-specific isoforms or isozymes: ‫مهم‬ - Proteins that differ somewhat in primary structure and properties from tissue to tissue, but that retain same function. - Notice that when we change amino acid with another amino acid with similar properties (e.g. hydrophobic to hydrophobic, polar to polar, positive to positive…), protein will usually retain its function. ‫مهم جدا‬  We even can compare same protein across different species : - Very small differences in the same protein give every species its distinct characteristics. - We study other organism, because same mechanisms and principles apply across species.  Importance of primary structure Sickle cell hemoglobin (HbS)  Primary structure of a protein determines other levels of structure : - It provides the built-in information to determine other levels of structure. - A single amino acid substitution in primary structure will result in change in secondary, tertiary and quaternary structure → give rise to a malfunctioning protein.  An example is Sickle-cell anemia : ‫فقر الدم المنجلي‬ - It is caused by a change of a single amino acid (Glu → Val) in 6th position of β-globin in hemoglobin. - This change is due to an inherited single nucleotide change in DNA. - The mutation results in: 1) Aggregates of millions of hemoglobin molecules inside RBCs. 2) Deformation of RBCs (become sickled cells like a crescent ‫ )مثل الهالل‬which makes them unable to carry oxygen efficiently. 3) Aggregation and clotting of RBCs in blood vessels and tissues.