BCH1002 Biochemical Aspects of Health and Disease

1. BCH1002Biochemical Aspects of Health and Disease BIOMOLECULES AND METABOLISM2. From Systems to Cells and Biomolecules Prof. K. M. Chan Dept. of Biochemistry Chinese University Rm 513B, Basic Medical Sciences Building Tel: 3163-4420; Email: kingchan@cuhk.edu.hk; chankingming@gmail.com Reference: On-Line Biology Book http://www.estrellamountain.edu/faculty/farabee/biobk/biobooktoc.html

2. Contents: • Biological Systems • Organizations of Cells • Organelles and their functions • Nucleic Acids: DNA and RNA • Proteins and amino acids • Sugars and carbohydrates • Fatty acids, lipids & membranes • Conclusions and revision exercises.

3. 2.1 From Systems to Cells • Systems> Organs> Tissues> Cells. • Various systems for specific life functions: Reproductive, cardiovascular (circulation), respiratory, nervous, digestive, endocrine, immune, etc. • Organs: specialized tissues for a unique function are to form an organ, e.g. pancreas, liver, lung, heart, intestine, brain, etc. • Tissues: Different cell-types for one unique task, e.g. pituitary gland to produce hormones, muscles (skeletal, cardiac, smooth muscles) to control movement, etc. • Cells: Specific cell type for one sole function, e.g. somatotroph is for growth hormone production and secretion, βcells in pancreas for insulin production and secretion.

5. Human Body Systems A Pathfinder for science students; visit this site to view different systems:http://pio.wsd.wednet.edu/library/body/bodypf.html The Endocrine System Hormone receptor http://www.estrellamountain.edu/faculty/farabee/biobk/BioBookENDOCR.html

6. The Reproductive Systems http://www.estrellamountain.edu/faculty/farabee/biobk/BioBookREPROD.html

7. Neuron Bone http://www.emc.maricopa.edu/faculty/farabee/biobk/BioBookAnimalTS.html#Muscle%20Tissue Skeletal Muscle

8. 2.2 Molecular Organization of Cells • Cells are basic units of function and structure • Robert Hooke (1665) first used cells (tiny boxes) to describe the structure which he observed inside a slice of cork under a microscope. • All cells are now known to be enclosed by a membrane that control the materials to go in and out of the cells. • All cells contain DNA as their genetic materials. Viruses could use either RNA or DNA as their genetic materials.

9. http://www.estrellamountain.edu/faculty/farabee/biobk/BioBookCELL2.htmlhttp://www.estrellamountain.edu/faculty/farabee/biobk/BioBookCELL2.html

10. Size Range of Cells • Small molecules (chemicals) < 1 nm (10-9 m) • Proteins and lipids- biomolecules < 10 nm • Ribosomes= 20 nm; viruses < 100 nm • Bacteria nm, 1 µm to 10 µm (10-6 m) • Mitochondrion, 2 µm; Nucleus, 7 µm • Plant and animal cells: 10-100 µm • Frog egg, 2 mm; salmon egg, 6 mm • Chicken egg, 4-5 cm • Length of muscle and nerve cells: 50-70 cm.

11. Bacterial cells with thick cell wall made of carbohydrates, could be differentiated as Gram + and – cells by Gram’s stain. Mesosome is infolding of membrane for specialized functions Cell wall Capsule http://www.bact.wisc.edu/themicrobialworld/structure.html Mesosome Membrane Nucleoid (DNA) region Ribosomes Contains lipo- polysaccharides (LPS) which are endotoxins of animals; but Gram-neg cells are sensitive to pennicillin and lysozyme. Flagella

12. Prokaryotic cells Small (1-5 μm diameter) No intracellular membrane organelles. Nucleoid with circular genomic DNA in cytoplasm, transcription and translation occur together. Often with cell wall for protection. E.g. bacterial cells Eukaryotic cells Larger and more complex cell (10 – 100 μm in length) Membraneous organelles for specific functions, e.g. mitochondria for respiration. Nucleus contains nuclear membrane, and genetic materials of DNA. After transcription, mRNA is processed and sent out of the nucleus for translation. Intracellular cytoskeletons. E.g. human, animal, plant and yeast cells. 2.2.1 Compare and contrast prokaryotic cells and eukaryotic cells

13. Compare (both are) Eukaryotic cells Contain organelles, for specific function Contain mitochondria Cytoskeleton (protein molecules) to hold up shape and structures Nucleolus in nucleus for rRNA synthesis Both contain rough endoplasmic reticulum and rough endoplasmic reticulum 2.2.2 Animal cells and plant cells

14. Contrast Plants cells and fungal cells have cell wall which is made of carbohydrate. Plant cells have mitochondria and chloroplast is only found in plants Vacuole and storage granules in plant cells for water control and nutrient storage Animal cells have lysosomes for digestion, microtubules, microfilaments, and centrioles for intracellular transport and cell division. Animal cells and plant cells

15. Adapted from Molecular Expression (http://micro.magnet.fsu.edu/cells/plantcell.html)

16. Mitosis and cell cycle http://teachline.ls.huji.ac.il/72373/substance_x/cell-cycle2.jpe http://www.cancerquest.org/index.cfm?page=58

17. Differentiation Adapted from the Merck Manuals On-line Medical Library: http://www.merck.com/mmhe/sec01/ch001/ch001b.html#sec01-ch001-ch001b-6

18. 2.3 Organelles and their functions • Cell is the basic unit of life defined by cell boundary set with membrane • In eukaryotic cells, membranes also define cellular organelles to carry out different specific functions at a well defined location: compartmentation. • Membrane is a selectively permeablebarrier between the cell and the external environment. • The lipid bilayer structure defines inner membrane and outer membrane sides. Two sides of the membrane show different properties. • Homeostasis- selective permeability allows the cell to maintain a constant internal environment. • Each organelles carry out a specific function

19. 2.3.1 The Nucleus • Bound by double membrane envelope that continues with endoplasmic reticulum. • There are nuclear pores to communicate with outside of the nucleus. “Gating” is necessary to ensure the various events impinging on gene transcription and cell signaling. • Luminal subunits in between the outer and inner membrane of the nucleus control the nuclear pores with a ring structure and nuclear cage. • Nucleolus is a structure inside the nucleus with chromatin actively transcribing ribosomal RNAs.

20. Nuclear translocator protein (vehicle) Architectures of nucleus Pore complex Outer membrane Inner membrane Luminal subunit Chromatin: DNA with histones and non-histone proteins Nuclear pores Ribosome: RNA translated to protein Inner membrane Outer Nuclear membrane Rough Endoplasmic Reticulum Nucleolus: ribosomal RNA genes being transcribed.

21. 2.3.2 Endoplasmic Reticulum (ER) • It spans the entire cell and attaches with numerous ribosomes for protein translation (Rough ER). • The proteins made in the ER are folded, may be glycosylated (post-translational modifications by adding sugars), and sorted to various parts of the cell. • ER also produces phospholipids (smooth ER). • It is also a depot of calcium ions (smooth ER) in the cell for controlling the signalling of cellular processes.

22. Rough ER dedicated for protein production http://cellbio.utmb.edu/cellbio/ribosome.htm#Ribosome-Endoplasmic%20Reticulum From RER to Golgi Apparatus http://cellbio.utmb.edu/cellbio/golgi.htm

23. 2.3.3 The Golgi Apparatus • Proteins made in the RER are sorted and processed in the Golgi apparatus. • They enter the Golgi at the cis face from vesicles formed at the ends of the ER. • In the cisternae, proteins are labelled (secondary modification), sorted and delivered to the trans face of the Golgi apparatus. http://sun.menloschool.org/~birchler/cells/animals/golgi/structure.html

24. Protein sorting (trafficking) • As vesicles formed in the Golgi, they are sent to proper locations for specific function to be carried out in other organelles or stored as secretory vesicles. • Proteins in vesicles would accumulate and wait for signal to be exported out by exocytosis (either regulated or constitutively secreted out of the cells. • Some proteins are delivered to and kept on cell membrane.

25. Rough Endoplasmic Reticulum and Golgi Apparatus Lysosome Early endosome Rough Endoplasmic Reticulum Engulfed materials Regulated secretory pathways Vesicle Trans Golgi apparatus Constitutive secretory pathway Cis-Golgi apparatus Golgi Apparatus Proteins are further modified and sorted in the Golgi apparatus

26. 2.3.4 The Lysosome • An intracellular digestive organ with acidic condition to digest and remove unwanted materials or break down materials for cellular uptake or re-use. • It contains digestive enzymes (primary lysosome). • After bud off from the Golgi, the enzymes may be released outside of the cellular membrane or fused with autophagic vesicles to form phagocytic vacuoles (secondary lysosomes) or endosomes). • Autophagy: degradation of intracellular components in lysosomes. • The debris could be discarded outside of the cell or kept as granules or recycle to cytoplasm.

27. 2.3.5 The MITOCHONDRION • Has double membrane: inner and outer. • The inner membrane fold to create the matrix space and inter-membrane space in between inner and outer membrane. • The matrix contains enzymes to oxidize metabolites • The inter-membrane has potential gradient generated by pumps at the inner membrane, energy released from the gradient produce ATP energy.

28. Outer membrane, with channel forming proteins but only permeable to 5 kDa molecules or less. Other enzymes on this membrane facilitate metabolism of lipid for use in matrix and transport of specially required protein in the mitochondrion. MAJOR FUNCTION OF MITOCHONDRION: OXIDATIVE PHOSPHORYLATION (oxidation of NADH to produce ATP) Matrix, internal space with enzymes for oxidation of pyruvate and fatty acids and for the citric acid cycle (Kreb’s cycle). Cristae in boxed region Intermembrane space, for enzymes using ATP passing out of the matrix to phosphorylate other nucleotides. Inner membrane, folded into numerous cristae to increase total surface area for electron-transport chain (respiratory enzymes), ATP synthase and transporters for metabolites going through the matrix.

29. 2.4 DNA and RNA Translation Transcription Protein RNA DNA Reverse Transcription The Central Dogma of Life Replication

30. Specific Cellular Function Transmission of Genetic Information Polypeptides RNA RNA polymerase Amino acids DNA Replication Semi-conservative Translation or Protein Synthesis Transcription or RNA synthesis

31. 2.4.1 Differences between DNA and RNA DNA, 2-deoxyribose sugar linked with A, G, C, or T bases. Usually double stranded For storage of genetic information RNA, ribose sugar linked with A, G, C, or U bases. Usually single stranded For gene expression, but some viruses use RNA as genetic materials PO4 PO4 5 5 CH2 CH2 O O Base 1 Base 4 2 3 3 OH OH OH H

32. 2.4.2 Formation of 5’ – 3’ Phosphodiester Bond PO4 RNA 5 CH2 DNA O O- 1 Base 5 4 2 OCH2 3 P O- Hydroxides can attack phospho-diester bond and cleave RNA, hence DNA is more stable. O 1 Base O- O OH 4 2 3 O P = O PO4 OH Phosphodiester bond linkage H O OH 5 5 CH2 CH2 O O 1 Base 1 Base 4 4 2 2 3 3 OH H OH H

33. 2.4.3 DNA Structure • Watson-Crick double helix (B-DNA) • Anti-parallel strands going opposite direction • A-T and G-C base pairing with hydrogen bonds (2 bonds for A=T, 3 bonds for G C) • A gene is a segment of DNA that controls and directs the synthesis of a polypeptide • Chromosome is a much larger structure with histones and other proteins packing up the DNA molecules with functional and structural DNAs.

34. Double helical structure of DNA and chromatin The DNA Double Helix 5’ 3’ Anti-parallel stranded with hydrogen bonds between bases to join the helixes together 11 nm in diameter H1 2 nm in diameter H2A H2B H3 H4 Histone protein 3’ 5’ Nucleosome core has 4 histones wrapped with one and three-quarters turns of DNA; H1 histone stays in between core; forming condensed fiber of 30 nm in chromatin structure. Each chromosome is 1 μm in diameter. Histones protect DNA from degradation by enzymes.

35. Bases: AT and GC pairing Pyrimidines: C, T NH Guanine H O N H C Purines: A, G HC C C C N C HC HN N Cytosine C N C N HN O H Hydrogen bond distance: 0.28 – 0.30 nm Adenine H CH3 HN N O H C Thymine C C C C N C HC HN N C N C N H O 1.08 nm

36. 2.4.4 Genetic Information can be decoded by transcription Eukaryotes: genes have introns and exons Prokaryotes: polycistronic, co-translation exon mRNA Introns, intervening region (uncoding) AAAAA RNA splicing AAAAA Mature messenger RNA AAAAA Nucleus Cytoplasm Translation in rough endoplasmic reticulum or polysomes area Proteins made Polycistronic: one mRNA codes for several proteins

37. 2.4.5 RNA, ribonucleic acids • All cellular RNAs are copied (transcribed) from a DNA template in the process of gene transcription. • Only messenger RNAs are further translated to make proteins. • Ribosomal RNAs and transfer RNAs do not make proteins directly but they form the translational machinery for protein synthesis. • Micro RNAs are small RNA molecules for gene regulation, they bind and cleave mRNAs to control gene expression.

38. Types and structures of RNAs O Uracil H H N • RNAs use uridine instead of thymine • Uracil could be produced by deamination of cytosine • RNAs are single stranded • They form secondary structures • They may form hairpins in G+C rich or A+U regions O N H PO4 Ribose OH OH U U U U U C G C C G C G C G C G C G AAUUUU UUUUUUU

39. RNAs have specific functions • Messenger RNAs carry information for amino acid sequence of a polypeptide chain to the ribosomes for translation for protein synthesis • Transfer RNAs are small molecules of 80 nucleotides that form three-dimensional structure and carry amino acids individually for translation • Ribosomal RNAs are part of the ribosomes with large size to form ribonuleoproteins for protein synthesis • Small nuclear RNAs and small cytoplasmic RNAs are for splicing, gene regulation, and control of gene expression.

40. Methionein 3’ ACC Amino acid attachment site 5’ microRNA to cleave RNA at specific sites 3’-OH Anticodon ACC UAC 3’ 5’ 5’ AUGAGC 3’ Codon 5’ Acceptor Stem TψC loop DHU loop Extra Arm Anticodon

41. 2.5 Proteins and Amino acids • Proteins are polypeptides. • Polypeptides is a polymer of amino acids connected in a unique sequence. • Amino acids are organic molecules possessing both carboxyl and amino groups with a side chain carrying different functional groups. • The carboxyl group of an amino acid is linked with the amino group of another amino acid and thus the amino acids are linked a polymer of polypeptide. • Different polypeptides can further linked together to make a larger protein complex.

42. 2.5.1 An overview of protein functions • Structural and support • Storage of amino acids or useful chemicals • Transporters • Hormones (messenger) • Receptors (membrane or cellular) • Contractile and movement • Enzymes • Antibodies for recognition • Transcription factors for DNA interactions 2.4 Proteins and Amino acids

43. At the center of the amino acid is an asymmetric carbon atom called alpha (α) carbon. The side-chain differs with amino acids. The R group may be as simple as hydrogen in glycine, or a carbon skeleton with functional groups attached as in glutamine. Peptide bond CO-NH is formed by dehydration. 2.5.2 Amino acids H O H C C N OH H R Amino Group Carboxyl end Side Chain • Polar, e.g. cysteine • Non-polar, e.g. glycine • Acidic, e.g. aspartic acid • Basic, e.g. lysine, arginine 2.4 Proteins and Amino acids

44. http://www.estrellamountain.edu/faculty/farabee/biobk/BioBookCHEM2.htmlhttp://www.estrellamountain.edu/faculty/farabee/biobk/BioBookCHEM2.html

45. http://www.estrellamountain.edu/faculty/farabee/biobk/BioBookCHEM2.htmlhttp://www.estrellamountain.edu/faculty/farabee/biobk/BioBookCHEM2.html

46. http://www.estrellamountain.edu/faculty/farabee/biobk/BioBookCHEM2.htmlhttp://www.estrellamountain.edu/faculty/farabee/biobk/BioBookCHEM2.html

47. 2.5.3 Synthesis of polypeptide stop 3’ 5 ‘ Start Termination Elongation 40 S 60 S Initiation In a polypeptide, the amino acids are joined with peptide bond between acid group of one amino acid and the amino group of another.

48. http://www.estrellamountain.edu/faculty/farabee/biobk/BioBookPROTSYn.htmlhttp://www.estrellamountain.edu/faculty/farabee/biobk/BioBookPROTSYn.html

49. Aminoacyl-tRNA synthetase reaction • The synthetase forms aminoacyl-AMP for the amino acid to be added to the 3’ end of the tRNA with the pyrophosphate. • Each amino acid would have their specific aminoacyl-tRNA synthetase. ribosome ATP mRNA 2.4 Proteins and Amino acids

50. A; anterior; P: posterior; E: exit sites P site Recognition of codon E site A site mRNA for translation Peptidyltransferase Peptide Bond Formation and moving onto P site, making the previous tRNA leave the P site and the A site for incoming tRNA-aa, until the stop codon appears. tRNA moves to E site to exit. Translocase 2.4 Proteins and Amino acids