Humboldt State University ® Department of Chemistry

Richard A. Paselk

Chem 438

Introductory Biochemistry

Spring 2007

Lecture Notes: 23 April

© R. Paselk 2006


Urea Cycle

Let's quickly look over the Kreb's Urea Cycle (this is his first cycle, the TCA cycle came later). Urea is synthesized from ammonia, carbon dioxide, and aspartate nitrogen using Kreb's Urea Cycle:

Note that two ATP's are required to synthesize carbamyl phosphate, the first to activate bicarbonate, the second to provide the activated phosphate on the carbamyl-P itself. Carbamyl-P is itself a high-energy compound with a mixed anhydride bond much like we saw in 1,3-bis PGA. The carbamyl group is then transferred onto ornithine which acts as a carrier for the growing urea molecule. Addition of the aspartate nitrogen requires two additional ATP equivalents to drive the condensation to argininosuccinate. Lysis results in the formation of a fumerate and arginine, which is then hydrolyzed to give our product, Urea, and regenerate the ornithine carrier.

Note a total of four ATP equivalents has been consumed. How many ATP equivalents are then required to convert the nitrogen of two amino acids into urea? The flow diagram supplied below may help in this calculation:

To solve this problem we can first note the four ATP equivalents consumed in the biosynthesis of one urea from two amino acids. But the fumerate produced must be taken back to regenerate the oxalacetate used to pick up the nitrogen from one amino acid):

fumerate + H2O malate; malate +NAD+ oxaloacetate + NADH + H+

This will provide an NADH via malate DH. This is equivalent to 2.5 ATP's, so we have -4 + 2.5 = -1.5. Next, while the second nitrogen enters via two transaminations through aspartate:

we also have to produce the ammonia from the first amino acid via glutamate using Glutamate DH,

This also produces an NADH, providing another 2.5 ATP's. thus the final tally is -4 + 2.5 + 2.5 = +1 ATP to produce one urea from two amino acid nitrogens. Of course for mammals this does not take into account the physiological costs of excreting the urea, which can be significant.



The enzymes involved in the conversion of amino acid nitrogen into urea occur in the cytosol and in the mitosol and involve three different catalytic systems: the Urea Cycle, The TCA cycle, and a transamination cycle, you might call them "Kreb's Tricycle." These interactions are shown in the figure in your packet (Kreb's Tricycle). Note the involvement of two antiports to move intermediates across the inner mitochondrial membrane: the malate: aspartate antiport, and the citrulline:ornithine antiport. Note also that nitrogen is incorporated in the mitosol (Transamination, Carbamoyl-P synthesis), whereas the final product, urea is released in the cytosol.


Tissue Distribution of Amino Acid Catabolism

Most amino acids are metabolized predominantly in the liver, but "alkyl" (branched chain) amino acids (val, leu, & ilu) are preferentially metabolized by skeletal muscle, whereas the "acyl" (asp, asn, glu, & gln) are metabolized in the intestinal mucosa. As an example let's look at the metabolism of protein after a one day fast (tissues need fuel and glucose). If we start with 1,000 mM of amino acids (equivalent to a "steak dinner" of 530 g of lean raw meat. (Data from McGilvery, Biochemistry: a Functional Approach, 1979, based on idealized calculations for amino acid distributions.):

Note that most of the alanine coming to the liver is from the muscle and intestine, produced from amino acid nitrogen and pyruvate to keep serum ammonia concentrations down. One of the most striking aspects of this chart is the difference in tissue usage of ATP (note that all calculations are based on 3 ATP/NADH and 2 ATP/FADH2). Both muscle and intestine derive significant energy the amino acids they breakdown. On the other hand, nearly all of the energy from the amino acids metabolized in the liver goes into glucose (very little ATP is made by the liver), which would go largely to the CNS (15,048 ATP @ 38 ATP's/Glucose, based on 3 ATP/NADH and 2 ATP/FADH2 used in the other calculations. If we use the values of 2.5 ATP/NADH and 1.5 ATP/FADH2, then we get 12,672 ATP @ 32 ATP/glucose)!


Alanine Cycle

Much of the nitrogen is carried between the tissues and the liver by the Alanine Cycle:

A similar, but more complex cycle involves glutamate/glutamine taking nitrogen from the muscle to the intestine where the glutamine is catabolized and the nitrogen goes to alanine which then goes to the liver. Glucose can then go to the muscle and be converted to glutamate via Glycolysis and the TCA cycle. We will not look at this more complex system.

Discussion of amino acid metabolism, lean meat, Lewis & Clark etc.

Catabolism of Amino Acid Carbon Skeletons

The catabolic breakdown of most of the amino acids is summarized in the Main Routes of Amino Acid Catabolism diagram in your packet. A couple of overview comments. Last time we looked at amino acids that are essential in mammals (cannot be synthesized in quantities necessary for good health). Amino acids can also be categorized as being glucogenic (can be used in Gluconeogenesis) or ketogenic (cannot be used in Gluconeogenesis). Most aa's can be at least partially used in glucose synthesis. For example ilu, tyr and phe are partially glucogenic and partially ketogenic(some carbons go to acetyl-CoA, while the rest go to TCA intermediates), while leu and lys are fully ketogenic.

We will begin by looking at the catabolism of amino acids by groups: 3-C (feed into pyruvate), 4-C (feed into oxalacetate), and 5-C (feed into glutamate).

3-C aa's: Ser and ala are converted in single step processes to pyruvate. Cys is converted after first oxidizing and removing sulfur as sulfate. (Threonine, glycine and part of tryptophan can also breakdown to pyruvate, but we will look at other paths.)

4-C aa's: Asn is hydrolyzed in one step to aspartate, which in turn is transaminated in one step to oxalacetate. Threonine feeds into the TCA cycle through succinyl-CoA instead of oxalacetate. Thr is first deaminated via a dehydratase as seen earlier, then decarboxylated by Pyruvate DH Complex to give propionyl-CoA, which is then transformed via a series of steps to give succinyl-CoA.


Propionyl-CoA metabolism: propionyl-CoA is an intermediate in the catabolism of a number of amino acids, as well as in the breakdown of odd-chain fatty acids. Propionyl-CoA (3-C) enters the TCA Cycle at succinyl-CoA (4-C), thus another carbon must be added to bring it into mainstream metabolism. A biotin-dependent carboxylase adds carbon dioxide at the cost of one ATP to give D-methylmalonyl-CoA. D-methylmalonyl-CoA is then racemized to L-methylmalonyl-CoA. Methylmalonyl-CoA is a branched-chain, whereas succinyl-CoA is straight-chain: the carboxyl group and a hydrogen must be exchanged. This exchange requires C-C bond-breaking and making, a process apparently involving a Co-C bond intermediate. The cobalamin cofactor derived from Vit B12 is used in catalyzing this reaction.

5-C aa's: Five aa's feed into glutamate which in turns feeds into the TCA cycle at 2-oxo-glutarate.

Branched chain amino acids: valine (val), leucine (leu), and isoleucine (ilu). The metabolism of each of these three amino acids begins with the same theme: transaminase; DH Complex; beta-oxidation. Due to the irreversible nature of the DH Complex all three are essential.

Pathway Diagrams

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Last modified 23 April 2007