BMB 401 Lecture 33 Transcript 1 Biochemistry 401 Lecture 33 Today we re going to talk about nucleotide function structure and synthesis We re going to talk about the de novo pathway and the salvage pathway So let s get started Let s begin by talking about nucleotide function Nucleotides are precursors for nucleic acid synthesis DNA and RNA and this is what we primarily think about what we think about nucleotides but they re also used for energy sources ATP is a prime energy source for biological processes such as molecular motors and active transport of solutes across membranes GTP is also sometimes used as an energy source Nucleotides can also activate biomolecules for synthetic reactions and we ve seen this previously for instance UTP activating glucose and CTP activating alcohol head groups and we ve also seen ATP activating many biomolecules Nucleotides and their derivatives are also important parts of many of many second messenger systems such as cyclic AMP and cyclic GMP Finally ATP is a source of phosphate for kinase reactions such as the ones that are involved in regulation We saw this in glycogen synthesis and degradation and also in glycolysis and so nucleotides serve many diverse functions So now let s talk about structure We re going to talk about structure of nucleosides first Nucleosides are comprised of two things sugars and nitrogenous bases Ribonucleosides are comprised of a ribose sugar and a base and deoxyribonucleosides are comprised of a deoxyribose sugar and a base and the difference is shown here We have adenosine that s shown in the top figure and deoxyadenosine that s shown in the bottom figure The difference between the two is that deoxyadenosine is reduced at the two prime position and the arrow is pointing to this position Now the bases can either be purines or pyrimidines and we ll see that in just a minute Nucleotides are nucleosides plus 1 2 or 3 phosphates In the top position we see adenosine a nucleoside and in the bottom position we see adenosine monophosphate AMP This is also known as adenylate In this slide we see the nomenclature of bases nucleosides and nucleotides It s important that you know the name of the base the name of the ribonucleoside or deoxyribonucleoside and the name of the ribonucleotide or deoxyribonucleotide BMB 401 Lecture 33 Transcript 2 And so it s important that you know the difference between adenosine and adenylate for instance Nitrogenous bases fit into two major categories Purines consist of adenine and guanine and if you ll notice these are structures made up of two rings Pyrimidines have one ring and there are three of them Cytosine uracil and thymine To remember this here are some mnemonics pure as gold for purines A and G cut pie C U T are all pyrimidines I hope this helps So now let s look at nucleotide synthesis There are two major pathways The first is the salvage pathway in which we use recycled bases and attach them to an activated ribose platform PRPP which is 5 phosphoribosyl 1 pyrophosphate The other way is the de novo pathway and this is making bases from scratch We start with an activated ribose again PRPP and we add amino acids ATP carbon dioxide and other things to make the finished nucleotide So in the salvage pathway 5 phosphoribosyl 1 pyrophosphate plus guanine for instance yields guanosine monophosphate and pyrophosphate and so this is driven forward by the pyrophosphate release and subsequent hydrolysis to two molecules of inorganic phosphate The de novo pathway is little more complex and pyrimidines and purines use two different strategies Pyrimidine rings are simple they re formed first and they re brought to the activated ribose platform PRPP and the subsequent progression involving several steps is first to make UTP and then to make CTP These are both ribonucleotides that can be used to make RNA Then UTP is used to make TMP and CTP is used to make dCTP These are both deoxyribonucleotides And so what we do first is we make the base first we attach it to PRPP and then we make ribonucleotides We use the ribonucleotides as substrates to make deoxyribonucleotides Now purines are much more complex They involve two rings These are made directly on the PRPP platform first to make the intermediate IMP then to make ATP and GTP So we re making ribonucleotides first and then to dATP and dGTP to make deoxyribonucleotides Purine rings are formed piece by piece right on the activated ribose sugar We re going to go over the primitive de novo pathway first as an overview and then we re going to go through it bit by bit We re going to do this with the pyrimidine de novo pathway but were not going to do the same thing with the BMB 401 Lecture 33 Transcript 3 purine de novo pathway So let s get started with this First of all we re going to synthesize an intermediate called orotate This is comprised of carbamoyl phosphate and aspartate We re then going to form orotidylate This occurs by attaching orotate to PRPP Then were going to form UMP uridylate This happens through a decarboxylation of orotidylate and then we re going to phosphorylate UMP to make UTP and then were going to aminate UTP to make CTP and these are all ribonucleotides The next that s going to happen we re going to make TPP and dCTP We re going to start with orotate synthesis So the first thing we re going to do is we re going to make orotate and it looks like this The green portions come from aspartate and the blue and black portions come from carbamoyl phosphate Now that s a name that should seem familiar to you This is the synthesis of carbamoyl phosphate The process is essentially the same as what we saw when we looked at the urea cycle Carbamoyl phosphate synthetase II actually catalyzes the rate limiting step in pyrimidine de novo synthesis and so it s really important that you understand this process This is the cytosolic enzyme that synthesizes carbamoyl phosphate Do you remember where the other carbamoyl phosphate synthetase was The one that s involved in urea synthesis is actually in the mitochondrion This one that s involved in nucleotide synthesis is found in the cytosol And so just like before we re going to start with bicarbonate We re going to phosphorylate it to make carboxy phosphate We re then going to aminate it to form carbamic acid then another phosphorylation to yield carbamoyl phosphate And so this is an activated intermediate And this is where the difference comes in If you ll remember in urea synthesis the amino group came
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