BMB 462 Exam 3 Study Guide Lectures 17 26 Lecture 17 Recognize the 5 bases and their associated nucleosides structures and physical chemical properties Nucleotides carry information and energy in the cell Nucleosides have a pentose sugar and a base attached If there is also a phosphate on the 5 atom they re called nucleotides can have up to 3 phosphates The base is flat while the sugar is not The flatness allows the bases to stack in the core of the DNA helix and that makes the DNA very strong The faces of the base are hydrophobic making hydrophobic stacking structure The edges are polar which is what allows you to make hydrogen bonds and make base pairing to other bases Depending on what base you have you can make different hydrogen bonds Use nucleotide nomenclature Base Nucleoside Nucleotide Nucleic Acid Purines Adenine Guanine Pyrimidines Cytosine Thymine Uracil Adenosine Deoxyadenosine Guanosine Deoxyguanosine Adenylate Deoxyadenylate Guanylate Deoxyguanylate RNA DNA RNA DNA Cytidine Deoxycytidine Thymidine Deoxythymidine Uridine Cytidylate Deoxycytidylate Thymidylate Deoxythymidylate Uridylate RNA DNA DNA RNA List nucleotide Properties Absorption Spectra All bases absorb light at 260nm this is due to the conjugated double bond in the bases which absorb light pH and tautomerization pH can change the form of uracil When pH is low uracil switches from lactam to lactim formation The Hydrogen moves from the N to the O on atom 2 This changes the hydrogen bonding In lactam N is the donor and O is the acceptor in lactim N becomes the acceptor H bonding and base pairing in DNA Because information is stored in the hydrogen bonding of DNA it is important to have the right pH b c changing pH changes H bonding and you will lose the information AT has 2 H bonds and GC has 3 so GC is slightly more stable Otherwise they are almost identical structurally The H bonding neutralizes the charge inside the DNA so the inside of the DNA becomes hydrophobic Then bases stack on each other and create a hydrophobic core that is very stable The outside of the DNA is hydrophilic List nucleotide Functions Building Blocks for DNA and RNA DNA transmits information from a parent cell to both daughter cells Information storage and transfer Information is stored in DNA and that is replicated and transferred through transcription into RNA RNA is then translated into proteins Energy Carriers Ribo nucleotides can also be used as energy carriers Mostly true for ATP and GTP So they can be building blocks or energy carriers Phosphoanhydride bonds are very high in energy so the cell to drive a lot of reactions uses them Coenzyme handles i e CoA NAD and FAD all have nucleotides included in them Signaling Molecules i e cAMP It s a signaling molecule inside the cell that acts as a secondary messenger Primary signals i e hormones are usually outside of the cell Compare and contrast salvage de novo purine synthesis and de novo pyrimidine synthesis Purines Pyrimidines Salvage Use PRPP Synthesis starts with Use PRPP Synthesis starts with Bases are constructed from PRPP and atoms are added to build PRPP and atoms are added to build intermediates produced in the base the base the degradative pathway of Atom sources for bases amino Atom sources for bases amino nucleotides acids CO2 and THF acids CO2 and THF Useful for cells that can t The bases are built attached to Orotate is first built and attached to do de novo synthesis ribose PRPP Orotate is then attached to Purines phosphoribosyl ribose and then is converted by transferases add PRPP to Intermediate Inosotate phosphoribosyl transferase into 2 bases Branched pathway Inosotate pyrimidines Pyrimidines Uridine either makes AMP or GMP Intermediate Orotate phosphorylase adds ribose Regulation feedback inhibition of Linear pathway orotate UTP CTP 1 phosphate to free uracil Regulation feedback inhibition of to make UMP the first step by IMP intermediate the first step by CTP AMP GMP Lecture 18 Describe the atom sources and reactions used in de novo nucleotide biosynthesis Sources of atoms for de novo synthesis a Purine i Nitrogen 3 nitrogen atoms come from Glutamine and aspartate amidotransferases the fourth is from glycine ii Carbon THF in the form of formate THF is common source of Carbon and more rarely CO2 can be a carbon source iii Glycine donates part of backbone b Pyrimidine i Aspartate donates part of backbone and the rest comes from carbamoyl phosphate carbamoyl phosphate comes from carbamoyl synthase c PRPP aka Phosphoribosylpyrophosphate Comes from pentose phosphate pathway which produces ribose 5 phosphate II Reactions a Phosphoribosyltransferases Attach a base to sugar to PRPP Explain the interconversion of nucleotide mono di and tri phosphates The enzyme that converts NMP to NDP nucleoside monophosphate kinases family of 4 enzymes The enzymes are specific for the base so there s one for A G C and U but not specific for the sugars can convert ribose and deoxyribose They use ATP to attach phosphate i e adenylate kinase uses ATP to convert AMP to ADP In this instance you actually make two ADPs because the ATP is converted to ADP as well The ATP then has to be regenerated Nucleoside Diphosphate Kinase converts all the different dinucleotides to trinucleotides ATP is again usually the donor and is converted to ADP Describe reduction of ribonucleotides to deoxyribonucleotides To make DNA the cell needs to convert all NDPs to dNDPs This is done by ribonucleotide reductase which uses e from NADPH to reduce the NDPs the e are transferred by thioredoxin Radical e are used to catalyze the reaction Ribonucleotide reductase reduces the 2 C by attacking 3 C and making it more reactive The enzyme has 2 subunits and subunit one has reactive thio groups attached to serine The hydrogen atoms come from NADPH and therefor need to be regenerated An active site radical on the R2 subunit is what causes the ribonucleotide radical to form on the 3 carbon That radical is what allows the chemistry to occur on the 2 C The radical allows the 2 hydroxyl to attack the thio group and protonate the 2 hydroxyl The protonated hydroxyl can then leave as water When water leaves it forms a carbocation The radical stabilizes the carbocation and the other SH group can now react with the 2 and reduce it Describe dNTP synthesis Ribonucleotide Reductase is a dimer so it contains 2 catalytic subunits and 2 active sites Chemistry can happen in both sites The two regulatory sites determine what is converted
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