Glycoproteins and Biogenic Amines - PDF

XY1110: BSc Medical Sciences Protein Function 2 Glycoproteins and Biogenic Amines Dr Peter Oatley [email protected]  Glycoproteins & Glycosamin...

XY1110: BSc Medical Sciences Protein Function 2 Glycoproteins and Biogenic Amines Dr Peter Oatley [email protected]  Glycoproteins & Glycosaminoglycans Lecture Layout Glycoproteins Glycosaminoglycans - Structure - Structure - - Amino Sugars - Function - N- or O- Linked - Proteoglycans - N-Linked - Types - Assembly & Modification - Function - Degradation - Degradation - Disease - Disease  Glycoproteins & Glycosaminoglycans Learning objectives Understand the functions of protein glycosylation Describe the differences between N- and O- linked glycosylation Know the basic structures of Gycosaminoglycans (GAGs) and proteoglycans Understand how disfunction in lysosomal targeting and degradation of glycoproteins can lead to disease Do you remember the structure of the plasma membrane? Great – please draw it with all the bits in ☺  Glycoproteins & Glycosaminoglycans Glycoproteins Glycoproteins - Proteins covalently linked to oligosaccharides (or Polysaccharides) - 10-15% oligosaccharide in weight  Glycoproteins & Glycosaminoglycans Glycoproteins Amino Sugars - Sugar + Amino and acetate groups Glucose signals carbohydrate availability  Glycoproteins & Glycosaminoglycans Glycosylation - N- or O- Linked N-Linked Glycosylation - N-acetylglucosamine (GlcNAc) - Amide nitrogen of an asparagine (Asn) - Consensus site: Asn-X-Ser/Thr - Glycosylated in the ER lumen  Glycoproteins & Glycosaminoglycans Glycosylation - N- or O- Linked O-Linked Glycosylation - N-acetylgalactosamine (GalNAc) + Others - Hydroxyl group of serine (Ser) or threonine (Thr) - Glycosylated in the Golgi lumen (via Sugar-Nucleotides)  Glycoproteins & Glycosaminoglycans N-Linked Glycosylation - Assembly & Modification Dolichol Phosphate - Spans the bilayer - Binds activated sugars - Enables “flipping” Oligosaccharyltransferase - Glycosylates proteins in the ER lumen….  Glycoproteins & Glycosaminoglycans N-Linked Glycosylation - Assembly & Modification  Glycoproteins & Glycosaminoglycans Glycoproteins - Degradation The Lysosome Stages of glycoprotein degradation 1) Endocytosis to Lysosome - From the cell surface 2) Protein Degrades - Proteases 3) Remove Sugars Asn - lysosomal glycosidases Just be aware there are different types and they are broken down in order  Glycoproteins & Glycosaminoglycans Glycoproteins - Degradation The Lysosome Disease Lysosomal hydrolases deficiencies → Lysosomal storage diseases - Undigested substrates in lysosomes Note: Defects often lead to nervous system defects - Change cell signalling - Decrease mitochondrial function → low [ATP]  Glycoproteins & Glycosaminoglycans Glycoproteins - Degradation Inclusion-cell disease (I-cell disease) Recessive mutation –rare - Hydrolytic enzymes missing - Fail to sort properly in the Golgi apparatus → Lysosomal hydrolases found in the blood Symptoms - Very severe Birth - Small, weak muscle tone Often: bone abnormalities - rounded upper back, clubfeet, dislocated hips Development - High mortality, Slow growth, stops at 2 years - Delayed motor skills and speech "coarse“ facial features and overgrowth of the gums  Glycoproteins & Glycosaminoglycans Glycosaminoglycans (GAGs) Glycosaminoglycans - Contain unbranched polysaccharides - With amino sugars and Acidic disaccharide repeats - Long and linear Example of a GAG acidic disaccharide repeat:  Glycoproteins & Glycosaminoglycans Glycosaminoglycans (GAGs) Glycosaminoglycans - Contain unbranched polysaccharides - With amino sugars and Acidic disaccharide repeats - Long and linear Example: Hyaluronan - A free GAG - ~25,000 disaccharide units - spun out directly from the cell surface - Large volume Space filler Compressive forces  Glycoproteins & Glycosaminoglycans Glycosaminoglycans (GAGs) Proteoglycans - Glycoproteins with GAGs - Up to 95% GAG in weight Biochemistry 9th Ed  Glycoproteins & Glycosaminoglycans Glycosaminoglycans (GAGs) Proteoglycans - Glycoproteins with GAGs - Up to 95% GAG in weight GAG Assembly on Protein 1) Tetrasaccharide added at serine in Golgi lumen 3) Repeats added one sugar at a time  Glycoproteins & Glycosaminoglycans Glycosaminoglycans (GAGs) Common Proteoglycan structures Proteoglycans - Glycoproteins with GAGs - Up to 95% GAG in weight Function: Lubricants in connective tissue Cell adhesion to extracellular matrix (ECM) Bind cell proliferation factors  Glycoproteins & Glycosaminoglycans Glycosaminoglycans - Degradation The Lysosome Stages of glycosaminoglycans degradation 1) Endocytosis 2) Remove Sugars - Lysosomal glycosidases - Last moiety added is first to be removed  Glycoproteins & Glycosaminoglycans Glycosaminoglycans - Degradation Hurler Syndrome Symptoms Genetic - Develop in first year of life or early childhood → GAGs not degraded - Large head (macrocephaly) and tongue → Enlarged lysosomes - Build-up of fluid in the brain → Excess GAGs stored in facial soft tissue - "coarse" facial features - Heart valve abnormalities - Enlarged liver, spleen - Decline in intellectual function Often - clouding of (cornea) - short stature - joint deformities - High morbidity in late childhood  Glycoproteins & Glycosaminoglycans Summary Glycoproteins Glycosaminoglycans - Function - cell surface, folding, resistance - Structure: amino sugars and Acidic - Predictions disaccharide repeats - Amino Sugars Long and linier - N- (Asn, GlcNAc) - Function - connective tissue - O- (Ser/Thr, GalNAc) - Proteoglycans - N-Linked - Types - Assembly & Modification - Function Dolichol Phosphate, Sugar- Lubrication/connective tissue Nucleotides - Assembly - - Degradation - lysosome Tetrasaccharide then single - Disease - I-cell disease - Degradation - last in 1st out - Disease - Hurler disease Questions?  Biogenic Amines Learning objectives Describe a biogenic amine and the various classes Understand how biogenic amines are synthesised and the functions of products Identify co-substrates and cofactors required for synthesis Know how biogenic amines are degraded and related disorders  Biogenic Amines Erdag et al (2018) Biogenic Amines, Ch1 Amino Acids - Other Uses Biogenic Amines Histamine - Small organic nitrogen containing molecules - Synthesised from amino acids Classes Tryptamine 1) Aromatic and heterocyclic Tyramine Serotonin  Biogenic Amines Erdag et al (2018) Biogenic Amines, Ch1 Amino Acids - Other Uses Biogenic Amines - Small organic nitrogen containing molecules - Synthesised from amino acids Classes 1) Aromatic and heterocyclic Putrescine 2) Aliphatic Mono-, di-, tri-, and poly-amines Cadaverine Spermidine Spermine Agmatine  Biogenic Amines Erdag et al (2018) Biogenic Amines, Ch1 Amino Acids - Other Uses Biogenic Amines - Small organic nitrogen containing molecules - Synthesised from amino acids Classes 1) Aromatic and heterocyclic 2) Aliphatic Mono-, di-, tri-, and poly-amines 3) Aliphatic volatile amines  Biogenic Amines Amino Acids - Other Uses Biogenic Amines - Small organic nitrogen containing molecules - Synthesised from amino acids Classes 1) Aromatic and heterocyclic 2) Aliphatic Mono-, di-, tri-, and poly-amines 3) Aliphatic volatile amines Functions - Cell membrane stabilization - Immune functions and inflammation mediators - Nucleic acid and protein synthesis - Hormones - Neural transmission  Biogenic Amines Biogenic Amine - Synthesis Many Reactions by…. 1) Decarboxylation of amino acids 2) Amination and transamination of aldehydes and ketones R1 R1 R2 R2 Erdag et al (2018) Biogenic Amines, Ch1  Biogenic Amines Biogenic Amine - Synthesis Amino Acid Biogenic Amine R Amino Acid Nucleotides for DNA/RNA  Biogenic Amines Biogenic Amine - Synthesis Amino Acid Biogenic Amine (From serine) Sphingolipids in membranes → cell signalling  Biogenic Amines Biogenic Amine - Synthesis Amino Acid Biogenic Amine Histidine Decarboxylase (From histidine)  Biogenic Amines Biogenic Amine - Synthesis Amino Acid Biogenic Amine (From tryptophan) Via Tryptophan decarboxylase Hormone AKA: Vitamin B3 - Relaxation - Coenzyme - Pleasure - Energy production - Memory  Biogenic Amines Biogenic Amine - Synthesis Amino Acid Biogenic Amine (From tyrosine) AKA: Adrenalin Thyroid gland hormone - Hormone → activated to triiodothyronine - Fight or Flight response (Liver/Kidneys) - Neurotransmitter - metabolic rate, muscle control, brain development, bone maintenance  Biogenic Amines Biogenic Amine - Synthesis UREA Cycle UREA Cycle Arginase in the UREA Cycle - A Ureohydrolase Poly-amine synthesis  Biogenic Amines Biogenic Amine - Synthesis Catecholamine Synthesis Biogenic monoamine neurotransmitters - Synthesized from amino acids 1) 5-hydroxytryptamine (5-HT) - hydroxylation and then decarboxylation of tryptophan 2) Histamine - decarboxylation of histidine 3x Catecholamines: - Tyrosine hydroxylated to Dihydroxyphenylalanine (DOPA) 3) Dopamine - decarboxylated DOPA 4) Norepinephrine (noradrenaline) - β-hydroxylated dopamine 5) Epinephrine (adrenaline) - methylated noradrenaline  Catecholamine Synthesis Biogenic Amines Biogenic Amine - Synthesis Coenzyme/cofactors Rate limiting: Tyrosine hydroxylase Co-substrate - Oxygen Cofactor - tetrahydrobiopterin, Fe2+  Biogenic Amines Biogenic Amine - Degradation Amine oxidases - Degrade biogenic amines R-CH2NH2 + H2O + O2 → R-CHO + NH3 + H2O2 Key: Enzymes Monoamine oxidases (MAO) Catechol-O-methyltransferase (COMT) Cosubstrate S-adenosyl-L-methionine (SAM) Prosthetic group Flavin adenine dinucleotide (FAD) Reductases, Dehydrogenases, Methyltransferases  Biogenic Amines Biogenic Amine - Degradation Amine oxidases - Degrade biogenic amines R-CH2NH2 + H2O + O2 → R-CHO + NH3 + H2O2 Key: Monoamine oxidases (MAO) Aldehyde dehydrogenase Cosubstrate S-adenosyl-L-methionine (SAM) Prosthetic group Flavin adenine dinucleotide (FAD) Reductases, Dehydrogenases, Methyltransferases  Biogenic Amines Biogenic Amine - Degradation Amine oxidases - Degrade biogenic amines R-CH2NH2 + H2O + O2 → R-CHO + NH3 + H2O2 Example: Monoamine oxidases (MAO) - Family of enzymes - Deaminates neurotransmitters - Requires Prosthetic group Flavin adenine dinucleotide (FAD) Types MAO-A - Serotonin, melatonin, norepinephrine & epinephrine MAO-B - Phenethylamine and benzylamine Both MAO -A and -B - dopamine, tyramine, and tryptamine  Biogenic Amines Biogenic Amine - Degradation Deficiency Build-up of Biogenic Amines - From body - From food/gut bacteria Example 1: Vitamin B12 (Folate) deficiency - Required for FAD → Monoamine oxidases disfunction → Tiredness, disturbed vision, psychological problems Example 2: Monoamine oxidases A deficiency - Very rare, often males - Serotonin brain build-up → Loss of impulse control Avoid High tyramine food (Aged cheese) Example 3: Inflammatory bowel disease etc. - Increased [Monoamine oxidases] activity → Oxidative stress → vascular dysfunction  Biogenic Amines Summary Biogenic Amines Biogenic monoamine neurotransmitters - Synthesis - Coenzyme/cofactors - Classes 1) 5-HT - tryptophan 1) Aromatic and heterocyclic 2) Histamine - histidine 2) Aliphatic Mono-, di-, tri-, and poly-amines 3x Catecholamines - tyrosine -> DOPA 3) Aliphatic volatile amines 3) Dopamine - Functions 4) Norepinephrine (noradrenaline) Membrane, immune/inflamation, nucleic acid, 5) Epinephrine (adrenaline) Hormones, Neural transmission Biogenic Amines Synthesis - Degradation - Coenzyme/cofactors - Reactions Monoamine oxidases (MAO) Decarboxylation, Amination & transamination Catechol-O-methyltransferase (COMT) - Amino Acids and products Trp, Arg & Ornithine - Degradation deficiency Vitamina B12, Monoamine oxidases A, IBD  Glycoproteins & Glycosaminoglycans Recommended reading Biochemistry Molecular Biology of the Cell 9th Edition 6th Edition Chapter 11 Ch13: Page 716-720 Ch19 Page 1058-1061