Ch7-120215 - CHEM 350: Introduction to Biological Chemistry...

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Unformatted text preview: CHEM 350: Introduction to Biological Chemistry Brian Lee, Ph.D. Office: Neckers 146G or 324 Phone: 453-7186 Ho urs: 9:30am to 10:30am or by appointment Website: https:/ / Textbook (required, U.S. edition only) Fundamentals of Biochemistry, 3rd Ed., Voet, Voet & Pratt. Study Guide (recommended) Student Companion to Fundamentals of Biochemistry, 3rd Ed. Help Desk Tuesday 6:30 to 7:30 pm in Neckers 218 Thursday 5:00 to 6:00 pm in Neckers 410 Announcements Undergraduate Research Opportunities Research for credit (such as CHEM 396 or CHEM 496) Student worker ($8.00 per hour) ( Undergraduate Assistantships ( McNair Scholars Program ( REACH Awards Competition ( Summer Research Experiences for Undergraduates (REU) For other REU programs, search the National Science Foundation site: Students must contact the individual sites for information and application materials. NSF does not have application materials and does not select student participants. A contact person and contact information is listed for each site. Assignments Read Chapter 8 Carbohydrates Chapter 8 Problems Student Companion site for Voet, Voet & Pratt Second Midterm Exam, Wednesday February 29th Chapters 6 through 9 All exams are cumulative Help Desk Tuesday 6:30 to 7:30 pm in Neckers 218 Thursday 5:00 to 6:00 pm in Neckers 410 Allosteric interactions: binding of a ligand at one site affects ligand binding at another site. Cooperativity between binding sites is seen in the sigmoidal binding curve, which switches between low and high affinity states (T -> R) Binding capacity for oxygen increases with increased cooperativity. (Note the difference in affinity at low pO2 and high pO2) Models of Cooperative Binding (Allosterism) Concerted Model assumes an all-or-nothing transition between T and R states. Ligand binding in “lock-and-key” fashion. Sequential Model follows the “induced fit” hypothesis. Ligand binding can change the conformational state. T state R state HILL PLOT: The Hill equation can be used to graphically determine n (Hill constant or coefficient). Note pO2=P50 at log Y 1Y =0 n = 1 non-cooperative (myoglobin n = 1) n > 1 + cooperative (hemoglobin n 3) n < 1 - cooperative (very rare) log Y 1Y = n log pO 2 n log p n50 CO2 Transport and the Bohr Effect CO2 transport is coupled to O2 transport by the exchange of protons between carbonic anhydrase and hemoglobin. HbH+ + O2 <-> Hb-O2 + H+ HCO3- + H+ <-> CO2 + H2O Cooperative binding of oxygen and the Bohr Effect is not enough: Allosteric inhibition by 2,3-BPG increases stability of the T state to allow release of oxygen in tissue Increases p50 from 12 to 26 torr Heterotropic Allosteric Regulation Regulation of O2 binding by 2,3-bisphosphoglycerate (BPG), a heterotropic allosteric regulator. BPG reduces affinity for O2 particularly at lower pO2 in tissues. BPG increases the difference in affinity between lungs and tissue. This change in affinity is directly related to the amount of oxygen deliverable to tissues. HbBPG + O 2 BPG + HbO 2 BPG stabilizes the T state (a and b). BPG has less affinity for the R state (c), since the binding pocket is too narrow T state R state Positively charged residues in the cavity of the T state, interact with the negatively charged groups on BPG Fetal hemeglobin has a higher affinity for oxygen. Allows the fetus to obtain oxygen from the maternal blood circulating through the placenta. Fetal hemeglobin contains dimers, where the subunit replaces the subunit of normal Hb. subunit has a lower affinity for 2 3-BPG Ser143 Ser143 Allosteric Regulation of Oxygen Binding to Hemoglobin Concerted versus Sequential Binding - Oxygen binding to T state triggers conversion to R state - All subunits undergo T state to R state transition Heterotrophic inhibitors increase cooperativity - 2,3-biphosphoglycerate allosteric inhibitor - Bohr Effect – mediated by H+, CO2 and ClFetal Isoform of Hemoglobin - subunit replaces subunit - lower affinity for 2,3-biphosphoglycerate - sacrifices binding capacity for increased affinity Sickle cell anemia is a genetic disease that manifests in homozygous individuals (two copies of same gene defect, known as hemoglobin S). Erythrocytes are deformed when the deoxygenated form of hemoglobin S aggregates within the cytoplasm, giving “sickle” shaped cells. Hemoglobin S contains Valine rather than Glutamate at position 6 in each chain. Val6 can fit into a hydrophobic pocket on another deoxyhemoglobin. This hydrophobic pocket is not present in oxyhemoglobin S. Cross-linked deoxyhemoglobin S forms strands which crystallize into fiber structures or aggregates. Erythrocytes are deformed when the deoxygenated form of hemoglobin S aggregates within the cytoplasm, giving “sickle” shaped cells. Molecular motors: Actin and Myosin in Striated Muscle Striated pattern in muscle fibers of: dark A bands and light I bands A band - contains overlapping thick and thin fliaments. I band - contains only thin filaments. H band - contains only thick filament. Thick filaments consist of myosin. Thin filaments consist of actin. Myosin 6 polypeptide chains - two 220 kDa heavy chains - two 15 kDa essential light chains (ELC) - two 22 kDa regulatory light chains (RLC) Myosin Heavy chain C–term tails associate to form the thick filaments. Head groups of myosin form cross-links to thin filaments. Thick filaments Light meromyosin tails are all two alpha helices associated through a coiled coil interface. The S1 fragment of the head group contains both ATP hydrolysis activity and actin binding activity. Thin filaments of F-actin G-actin bound to ATP analog. ATP hydrolysis drives assembly ELC actin binding surface RLC ATP binding site Myosin head group cross-links with one subunit of F-actin Sliding filament model The force of muscle contraction is generated by thick and thin filaments sliding past each other. (Huxley & Niedergerke, 1954) Myosin “walks” along the actin filament, driven by ATP hydrolysis Myosin motor domain changes conformation leading to a 60° shift in orientation and a 100Å shift in position along the actin filament. ADP + VO43- is a mimic for ADP + Pi which is the “high energy” state. The “low energy” state is after the “power stroke” and release of ADP. Myosin “walking” Control of Muscle Contraction Nerve impulses release Ca2+ from sarcoplasmic reticulum. Ca2+ binds to troponin (TnC) which causes a change in conformation of the troponin-tropomyosin complex. In the absence of Ca2+, tropomyosin (Tm) blocks the binding sites for myosin in the actin filament. Thin filaments of F-actin G-actin bound to ATP analog ATP hydrolysis drives assembly Actin filaments in non-muscle cells mediate cell motility Microfilament treadmilling driven by ATP hydrolysis. Immune response: 1. Cellular immune system: • guards against virally infected cells, fungi, parasites and foreign tissue • mediated by T Lymphocytes or T cells (develop in the Thymus) 2. Humoral immune system: • most effective against bacterial infections • mediated by a diverse collection of related proteins known as antibodies or immunoglobulins • antibodies are produced by B lymphocytes or B cells (mature in Bone marrow). • (humor is an archaic term for fluid) Cellular Immune System, Part I: “ The Killer T Cells” 1) Macrophage engulfs pathogen 2) Pathogen digested by enzymes in lysosome 3) Fragments display on surface by Class I MHC fragment = antigen MHC = Major Histocompatibility Complex Cellular Immune System 4) Macrophage is now an antigen-presenting cell (APC). Class I MHC and antigen is recognized by T cell receptor on immature cytotoxic T cell (killer T cells) 5) APC and T cell release cytokine to stimulate growth Cellular Immune System 6) Cytokines (interleukin-1 and interleukin-2) stimulate T cell proliferation - new T cells recognize same antigen 7) T cells only release interleukin-2 when bound to APC, which limits immune response to pathogen Cellular Immune System 8) mature cytoxic T cells are targeted to antigen 9) all cells display Class I MHC with self and foreign antigens cells displaying foreign antigen are bound by cytotoxic T cells 10) cytotoxic T cells destroy infected host cells by apoptosis Cellular Immune System, Part II: “ The Helper T Cells” 1) Macrophages also display Class II MHC proteins 2) Pathogens are fragmented 3) Macrophages display Class II MHC and antigen 4) immature helper T cells bind Class II MHC and antigen Cellular Immune System 5) APC and helper T cell release cytokines 6) cytokines stimulate proliferation of helper T cells 7) interleukin-2 from helper T cells also stimulates proliferation of cytotoxic T memory cells Cellular Immune System 8) mature helper T cells can become long-lived helper T memory cells (11) 9) mature helper T cells also play a role in the humoral immune system by interaction with B cells Humoral Immune System 1) B cells display both immunoglobulins and Class II MHC 2) B cells engulf antigen recognized by immunoglobulin 3) antigen is fragmented by enzymes in lysosome 4) Class II MHC and antigen are displayed on surface Humoral Immune System 5) 6) 7) 8) Mature helper T cells recognize Class II MHC and antigen Interleukins stimulate B cell proliferation B cell division continues in the presence of helper T cells Most B cell progeny are plasma cells, which make large amounts of antigen specific immunoglobulins (antibodies) Humoral Immune System 9) Antibodies bind to available antigens and mark them for destruction by white blood cells (phagocytes) 10) B cells can also become long-lived B memory cells which allow for rapid response to later infection by the same pathogen (secondary immune response) Memory cells enable the secondary immune response ...
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This note was uploaded on 03/26/2012 for the course CHEM 350 taught by Professor Lee during the Spring '08 term at SIU Carbondale.

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