The Raw Materials of Biotechnology

1. The Raw Materials of Biotechnology

2. Learning Outcomes • Identify the levels of biological organization and explain their relationships • Describe cell structure and its significance in biotechnology research and product development • Discuss the types of organisms researched and the types of cells grown and studied in biotechnology facilities plus the product with which they are associated

3. Learning Outcomes • Distinguish between the cellular organization of prokaryotic and eukaryotic cells • List the 4 main classes of macromolecules and describe their structure and function • Define genetic engineering and identify products created with this technology • Explain the Central Dogma of Biology and its importance in genetic engineering

4. Vocabulary • Fluorometer • Organism • Cell • Escherichia coli • Multicellular • Cytology • Anatomy • Physiology • Respiration • Unicellular • Chlorophyll • Photosynthesis • Steroids • nucleotide • Tissue • Organ • Protein • Eukaryote • Protist • Organelles • Mitochondria • Sugar • Starch • Nucleic acid • Pancreas • Hormone • R group • ribonuicleic acid

5. Vocabulary • Chloroplast • Cytoplasm • Lysosome • Ribosome • Cell wall • Cellulose • Plasma membrane • Glucose • ATP • Nucleus • Chromosomes • Enzyme • Cellular respiration • Deoxyribose • Hydrophobic • Triglycerides • Ribose • Phospholipids • Pigments • mRNA • Amino acid • Polypeptide • Chinese hamster ovary cells • Vero cells • Prokaryote • Organic • Carbohydrates • Cytoskeleton • Monomer • Polymer • Monosaccharide • Disaccharide • Polysacccharide • Fructose • Sucrose • Lactose • Hydrophillic

6. Organisms & Their Components • To manufacture biotechnology products for medical, industrial, or agricultural applications, biotechnicians must work either directly or indirectly with organisms or their components • Entire multicellular organisms • Bacteria • Tissues/cells

7. Organisms & Their Components • Working in any area of biotechnology requires a thorough understanding of the characteristics of life and the structures that compose organisms • You must have at least a minimal understanding of • Biochemistry: organic molecules structure, function, & interactions • Cytology: branch of life science, which deals with the study of cells in terms of structure, function and chemistry • Anatomy: structure of living things and their components • Physiology: the processes and functions of living systems

8. The Living Condition • Biologists estimate that there are well in excess of 20million different species, with some estimates being as high as 150 million • All living things can be classified into one of the following categories

9. The Living Condition • Biologists estimate that there are well in excess of 20million different species, with some estimates being as high as 150 million • All living things can be classified into one of the following categories

10. Characteristics of Life • Growth & Development • Reproduction • Organized structure composed of 1 or more cells • Response to stimuli • Conversion of energy • Respiration • Metabolism • Catabolism • Anabolism

11. Here We Go Again! Cells & Their Parts • The cells of both unicellular and multicellular organisms are tiny microscopic factories that produce thousands of different molecules • Biotechnology companies exploit the biologiical manufacturing capabilities of cells and trick them into producing particular molecules in large quantities • These become biotechnology products • Organelles: a specialized subunit within a cell that has a specific function, and is usually separately enclosed within its own lipid bilayer • The name organelle comes from the idea that these structures are to cells what an organ is to the body

12. Eukaryotic Cell Structure

13. Organelles involved in Energy Production Mitochondria Chloroplast • A membrane bound organelle consisting of an outer membrane and a highly folded inner-membrane; it is responsible for generating large amounts of ATP through the process of aerobic cellular respiration • Found in all eukaryotic cells • ATP: adenosine triphosphate a nucleotide that serves as the main source of usable energy for ALL living things • A specialized organelle in plants and some protists that uses the energy from sun light to convert inorgaic molecules (CO2 & H2O) into energy storing organic molecules (glucose) through the process of photosynthesis

14. Organelles involved in Energy Production Mitochondria Chloroplast • A membrane bound organelle consisting of an outer membrane and a highly folded inner-membrane; it is responsible for generating large amounts of ATP through the process of aerobic cellular respiration • Found in all eukaryotic cells • ATP: adenosine triphosphate a nucleotide that serves as the main source of usable energy for ALL living things • A specialized organelle in plants and some protists that uses the energy from sun light to convert inorgaic molecules (CO2 & H2O) into energy storing organic molecules (glucose) through the process of photosynthesis

15. Nucleus: The Organelle that Stores the Hereditary Material Nuclear Envelope Nuclear Contents • a double lipid bilayer that encloses the genetic material in eukaryotic cells. • serves as the physical barrier, separating the contents of the nucleus from the cytosol • nuclear pores are inserted in the nuclear envelope, which facilitate and regulate the exchange of materials (proteins such as transcription factors, and RNA) between the nucleus and the cytoplasm. • The outer membrane is continuous with the rough endoplasmic reticulum • The outer and inner nuclear membrane are fused at the site of nuclear pore complexes. • DNA: a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms • main role is the long-term storage of information • Used as a template to make RNA through the priocess of transcription • Nucleolus: a non-membrane bound structurecomposed of proteins and nucleic acids found within the nucleus. • Ribosomal RNA (rRNA) is transcribed and assembled within the nucleolus, forming ribosomes

16. Nucleus: The Organelle that Stores the Hereditary Material Nuclear Envelope Nuclear Contents • a double lipid bilayer that encloses the genetic material in eukaryotic cells. • serves as the physical barrier, separating the contents of the nucleus from the cytosol • nuclear pores are inserted in the nuclear envelope, which facilitate and regulate the exchange of materials (proteins such as transcription factors, and RNA) between the nucleus and the cytoplasm. • The outer membrane is continuous with the rough endoplasmic reticulum • The outer and inner nuclear membrane are fused at the site of nuclear pore complexes. • DNA: a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms • main role is the long-term storage of information • Used as a template to make RNA through the priocess of transcription • Nucleolus: a non-membrane bound structurecomposed of proteins and nucleic acids found within the nucleus. • Ribosomal RNA (rRNA) is transcribed and assembled within the nucleolus, forming ribosomes

17. Nucleus: The Organelle that Stores the Hereditary Material Nuclear Envelope Nuclear Contents • a double lipid bilayer that encloses the genetic material in eukaryotic cells. • serves as the physical barrier, separating the contents of the nucleus from the cytosol • nuclear pores are inserted in the nuclear envelope, which facilitate and regulate the exchange of materials (proteins such as transcription factors, and RNA) between the nucleus and the cytoplasm. • The outer membrane is continuous with the rough endoplasmic reticulum • The outer and inner nuclear membrane are fused at the site of nuclear pore complexes. • DNA: a nucleic acid that contains the genetic instructions used in the development and functioning of all known living organisms • main role is the long-term storage of information • Used as a template to make RNA through the priocess of transcription • Nucleolus: a non-membrane bound structurecomposed of proteins and nucleic acids found within the nucleus. • Ribosomal RNA (rRNA) is transcribed and assembled within the nucleolus, forming ribosomes

18. Ribosomes: involved in Protein Synthesis • Non-membrane bound organelles composed of rRNA & protein that provide the appropriate microenvironment to catalyze peptide bond formation between amino acids based on the instructions it reads on an mRNA molecule through the process of translation • Chemistry Connect: Remember, all organic polymers are formed through condensation reactions, aka dehydration synthesis

19. Organelles involved in Trafficking Rough Endoplasmic Reticulum Golgi Apparatus • A highly folded lipid bilayer whose surface ois studded with protein-manufacturing ribosomes giving it a "rough" appearance • Ribosomes bound to the RER at any one time are not a stable part of this organelle's structure as ribosomes are constantly being bound and released from the membrane. • A ribosome only binds to the ER once it begins to synthesize a protein destined for the secretory pathway • The Golgi apparatus processes and packages macromolecules, such as proteins and lipids, after their synthesis and before they make their way to their destination; it is particularly important in the processing of proteins for secretion.

20. Organelles involved in Trafficking Rough Endoplasmic Reticulum Golgi Apparatus • A highly folded lipid bilayer whose surface is studded with protein-manufacturing ribosomes giving it a "rough" appearance • Ribosomes bound to the RER at any one time are not a stable part of this organelle's structure as ribosomes are constantly being bound and released from the membrane. • A ribosome only binds to the ER once it begins to synthesize a protein destined for the secretory pathway • The Golgi apparatus processes and packages macromolecules, such as proteins and lipids, after their synthesis and before they make their way to their destination; it is particularly important in the processing of proteins for secretion.

21. Organelles involved in Metabolism Smooth Endoplasmic Reticulum Lysosome • Functions in several metabolic processes, including synthesis of lipids and steroids, metabolism of carbohydrates, regulation of calcium concentration, drug detoxification, attachment of receptors on cell membrane proteins, and steroid metabolism. • It is connected to the nuclear envelope. • Lysosomes: contain acid enzymes to break up waste materials and cellular debris. • found in animal cells, while in yeast and plants the same roles are performed by lyticvacuoles. • digest excess or worn-out organelles, food particles, and engulfed viruses or bacteria • The membrane around a lysosome allows the digestive enzymes to work at the 4.5 pH they require.

22. Organelles involved in Metabolism Smooth Endoplasmic Reticulum Lysosome • Functions in several metabolic processes, including synthesis of lipids and steroids, metabolism of carbohydrates, regulation of calcium concentration, drug detoxification, attachment of receptors on cell membrane proteins, and steroid metabolism. • It is connected to the nuclear envelope. • Lysosomes: contain acid enzymes to break up waste materials and cellular debris. • found in animal cells, while in yeast and plants the same roles are performed by lytic vacuoles. • digest excess or worn-out organelles, food particles, and engulfed viruses or bacteria • The membrane around a lysosome allows the digestive enzymes to work at the 4.5 pH they require.

23. Metabolism: is the set of chemical reactions that happen in living organisms to maintain life Catabolism Anabolism • is the set of pathways that break down molecules into smaller units • These reactions release energy • the set of metabolic pathways that construct molecules from smaller units. • These reactions require energy Critical Thinking: Which organelles are catabolic and which are anabolic?

24. The Endomembrane System • Is composed of the different membranes that are suspended in the cytoplasm within a eukaryotic cell. • These membranes divide the cell into functional and structural compartments, or organelles. • In eukaryotes the organelles of the endomembrane system include: the nuclear envelope, the endoplasmic reticulum, the Golgi apparatus, lysosomes, vacuoles (plants), vesicles (all eukaryotes), and the cell membrane

26. Cellular Membranes • Acts as a semi-permeable barrier that regulates what enters or leaves the cell or organelle • They consist of phospholipids which spontaneously arrange so that the hydrophobic "tail" regions are shielded from the surrounding polar fluid, causing the more hydrophilic "head" regions to associate with the cytosolic and extracellular faces of the resulting bilayer.

27. Cellular Membranes • Acts as a semi-permeable barrier that regulates what enters or leaves the cell or organelle • They consist of phospholipids which spontaneously arrange so that the hydrophobic "tail" regions are shielded from the surrounding polar fluid, causing the more hydrophilic "head" regions to associate with the cytosolic and extracellular faces of the resulting bilayer.

28. Things in/on Cellular Membrane • Proteins: function in cellular communication, catalyzing reactions, cell, signaling, cell movement • 4 main types • Receptor proteins bind signal molecules and initiate signal transduction pathways • Enzymes: speed up chemical reactions • Channel proteins: open or close to let target molecules in or out in response to a signal • Marker proteins: specific for each cell type; also, help the immune system distinguish self from non-self • Cholesterol: establishes proper membrane permeability & fluidity

29. Cell Walls • All living things except animals and some protists have cell walls • Cell walls are semipermeable boundaries that are more rigid than, and surround the plasma membrane • provides structural support and protection, and also acts as a filtering mechanism • Keeps cells from bursting in hypotonic environments • Different organisms have cell walls made of different organic polymers • Plants: cellulose • Bacteria: peptidoglycan • Fungi (some): chitin

30. Cell Tissue  Organ  Organism • Depending on the type of cell hundreds of different molecules are being manuifactured at any given moment • Some molecules are unique to a specific type of cell, others are produced in all cells • Example: Pancreas organ of the digestive system that lies in your abdomen, behind your stomach,with 2 main functions: • producing digestive enzymes to break down food; and • producing the hormones insulin and glucagon to control sugar levels in your body.

31. Case in Point: Pancreas

32. A closer look at the BIG Picture • When you, the organism, eats a meal, your stomach (with the help of digestive enzymes from your pancreas and other organs) break down the complex carbohydrates into glucose • Glucose is absorbed through the intestines (small I think) into the blood stream • Rising glucose in the blood stream is detected by the pancreas as the glucose binds glucose receptors on pancreatic cells. This stimulates β-cells to release insulin

33. Insulin: pancreatic homeostasis • Insulin works by improving the uptake of glucose from the blood across cell membranes and into the cells of the body by binding to insulin receptors on cells • Specifically, when insulin binds the insulin receptor, the receptor protein transports phosphate groups from ATP to other proteins in the cell, initiating signal transduction pathways that open glucose channels allowing passive diffusion of glucose into the cell • Once in the cells, the glucose is used as the energy to fuel the cells doing their different jobs or is stored in the liver or muscle cells as glycogen. • This results in the glucose level of the blood dropping, which then triggers the pancreas to switch off the release of insulin

34. Critical Thinking • Insulin and glucagon are both hormones that work together to regulate blood glucose levels. Knowing how insulin works, and that glucagon's effect is opposite that of insulin, propose a mechanism by which glucagon may work • Hint: remember that unused glucose is converted to and stored in the form of glycogen in the pancreas

35. Glycogenolysis • Glucagon helps maintain the level of glucose in the blood. • Glucose is stored in the liver in the form of glycogen, which is a starch-like polymer chain made up of glucose molecules. • Liver cells (hepatocytes) have glucagon receptors. • When glucagon binds to the glucagon receptors, the liver cells convert the glycogen polymer into individual glucose molecules, and release them into the bloodstream, in a process known as glycogenolysis.

37. Why all this matters • The purpose of biotechnology is to increase quality and/ or quantity of life • Life is dependant on the bodies abilities to • Maintain homeostasis • Transform energy • Reproduce • Grow and develop • Respond to stimuli • Internal: for example a change in levels of blood sugar • External: for example a change in temperature Biotechnology focuses on making organisms better able to accomplish one or more of these tasks

38. Central Dogma • If the goal of biotechnology is to increase an organisms ability to accomplish one or more of the characteristics of life, then it is important to understand how the organism naturally carries these tasks out! • One of the most important aspects of understanding this is to understand the central dogma of biology

39. Chromosomes & Genes • We know chromosomes are in the nucleus • We know genes are on chromosomes, and are segments of DNA that code for protein • What we need to look at more specifically is HOW the DNA instructs your body to build a protein

40. Central Dogma: an Overview • DNA is made up of 2 strands of nucleotides that coil around each other forming a double helix • Each strand is composed of varying arrangements of 4 nucleotides • Adenine Cytosine Thymine Guanine • When genes are used to instruct the body to build a protein, they must be transcribed into a complementary sequence of messenger RNA • The mRNA transcript is made of nucleotides • Adenine Cytosine Uracil Guanine

41. Central Dogma: an Overview • This mRNA can leave the nucleus and be read by a ribosome which actually constructs the protein through the process of translation • In translation, the ribosome links amino acids together in the order specified by the sequence of nucleotides in the mRNA • These amino acids are held together by special covalent bonds called peptide bonds • There are 20 different amino acids • A chain of amino acids are called a polypeptide

42. Nucleus Ribosome in cytoplasm or on RER

43. Central Dogma • The process of Gene Expression is universally found in all cells • How the DNA code is rewritten into mRNA and then decoded into a protein • It is called “The Central Dogma of Biology” because it helps explain how virtually all molecules are made either • directly protein & nucleic acids • or indirectly carbohydrates and lipids because their synthesis is controlled by the proteins

44. Tying it together • DNA is used to make protein • Proteins do most of the work of the cell • The other organic molecules are important too, but because their production/ destruction is controlled by proteins we focus on the protein • The cell has specialized organelles, containing different collections of proteins, giving each organelle a unique function based on the proteins (and other organic molecules) in/on it • The organelles work together to help the cell carry out all of the characteristics of life

45. Tying it together • Not all cells, even within the same individual, are the same • Some cells become specialized to perform a specific function • β-cells: Secrete insulin • Rods: sense light • Cells are able to perform these specialized functions BECAUSE of the proteins they produce • Different types of cells produce different types/quantities of proteins at different times

46. Critical Thinking • If all cells within one organism have the same set of DNA (in other words, the DNA in your nerve cells is identical to the DNA in the epithelium of your little toe), how is it that all cells are not the same?

47. Cells Commonly Used in Biotechnology • In biotech applications, some cells are used more than others • Chinese Hamster Ovary (CHO)cells • Vero Cells: African green monkey kidney epithelial cells • HeLa cells: human epithelial cells • Fungal cells • Aspergillus • yeast • Prokaryotic cells • E. coli • Staphylococcus • Streptococcus In Other Words You need to know how to keep cells alive in culture, in order to do most biotechnology research

48. NOTE • On page 46 of your text book it claims that • “Due to their lack of mitochondria, bacteria conduct ONLY anaerobic respiration” • This is WRONG & the author is a MORON • There are SOME strictly anaerobic bacteria but MOST need or at least can survive in oxygen environments!

49. Read section 2.3 on your own! • Things to pay attention to • Carbohydrates • Glycogen: what is it • Structural polysaccharides of cell wall • How polysaccharides interefere with purification procedures • The text book reads “Cells break the bonds in glucose, releasing energy in a form that cells can use” why do I hate that definition • Figure 2.23 • lipids • why are lipids referred to as hydrocarbons • The 3 general groups of lipids and their functions • Triglycerides • Phospholipids • Steroids • Structure of a phospholipid

50. “DNA is the Flash, Proteins are the Cash!” • Water makes up 75% of a cells mass • Of 25% of dry mass, 75% of it is protein • In biotechnology, proteins are often the manufactured product • Often, 50-75% of a companies staff is devoted to protein research (proteomics)