Monday, July 11, 2011

Glycolysis (enzymes of the preparatory phase)

As I have already mentioned on other posts, glycolysis is composed by 10 biochemical reactions catalyzed by enzymes all different. Today I dedicate this post to some information on the enzymes of the preparatory phase.
1st enzyme - Hexokinase
This enzyme, present in all our cells, has different isoforms present in our body and is the first point of regulation of glycolysis. In general, regardless of the isoform considered, its mass is about 100kDa. It is an enzyme that can be structurally divided into two halves with plenty of homology, the N-terminal half and half C-terminal. Because of this characteristic, it is thought that the gene for this enzyme may have arisen by duplication of an ancestral gene. The 3D structure of hexokinase can be compared to the shell of a bivalve...
There are four major isoforms of hexokinase (I-IV), and the fourth may also be called glucocinase (or hexokinase D), and is found primarily in the liver. Glucocinase The kinetic properties and regulatory features significantly different from other isoforms. The hexokinase I-III have a very high affinity for glucose (Km for glucose is about 0.1 mM), and to a normal concentration of glucose (4-5 mM) the enzyme is saturated with substrate. That is, the amount of available substrate is sufficient for the enzyme to function at its maximum speed. On the other hand, glucocinase presents a much higher Km (10 mM), which means that under normal conditions the enzyme is far from saturated with substrate. Probably at this point you will ask: "What's the point of this? It should be much more advantageous to have an enzyme to function at its maximum speed!" The answer to this question is very simple ... The function of glucocinase is to produce glucose-6-P which is then diverted mainly to the synthesis of liver glycogen. Thus, it only makes sense we have a lot synthesize glycogen when glucose availability is high. Therefore, the glucocinase will only start operating at a higher speed if there is an increase in the substrate availability. In other words, unlike what happens with the other hexokinase, when higher the concentration of glucose increased the speed of action of glucocinase.

The main substrate of hexokinase is D-glucose, but can also use other substrates such as hexoses, such as D-fructose and D-mannose. However, the value of Km for these substrates is higher, ie, the enzyme can use them but has less affinity for the same. This situation occurs mainly caters for hexokinase I-III, and the glucocinase is more specific for glucose.

The mechanism of action of hexokinase is called the Random Bi Bi, in which the enzyme forms a ternary complex with glucose and the Mg2 +-ATP before the reaction occurs. It makes a catalysis by proximity effect.
2nd enzyme - Fosfohexose isomerase
This enzyme has an activity highly dependent on pH, suggesting a mechanism of action involving charged side chains of amino acids in its active center. In fact, the presence of a glutamate and a lysine in the active site of fosfohexose isomerase is essential for the catalytic activity of the same. This enzyme is highly steroespecific.

3rd enzyme - Phosphofructokinase-1 (PFK-1)
The PFK-1 is the second regulatory enzyme of glycolysis, and is its main point of regulation. Presents a certain analogy with hexokinase, because the reaction is identical to that catalyzes the first reaction of glycolysis. At the structural level, it presents as a homotetramer.

There is another PFK, the PFK-2, which does not act directly in glycolysis, but is central to its regulation, because it controls the levels of fructose-2 ,6-bisphosphate, an important activator of PFK-1! (I will soon put a post on the regulation of glycolysis ...)
4th enzyme - Aldolase
This enzyme is highly steroespecific. Presents three different isoforms (A, B and C), whose expression varies during the development of the organism. The major isoform in humans is the isoform A.

n glycolysis, the aldolase catalyzes a reaction known as retro-aldol condensation. There are two amino acid residues essential for the activity of the enzyme, a lysine and a cysteine.

5th enzyme – Triose phosphate isomerase
The triose phosphate isomerase appears as a homodimer. Each subunit has a barrel-shaped structure, composed of eight alpha helices and eight parallel beta sheets. It was the first enzyme discovered to exhibit this type of barrel alpha / beta. This enzyme has a high dependence on catalytic function of pH, which indicates that performs an acid-base catalysis. In fact, there are three amino acid residues essential for its activity, a glutamate, a histidine and lysine. These amino acid residues play a role towards the establishment of hydrogen bonds that stabilize the transition state. Additionally, the enzyme has a loop with 10 amino acid residues highly conserved. This loop is important to stabilize the enediol (intermediate reaction) formed during the catalytic activity of the enzyme.

Triose phosphate isomerase is often mentioned as a case of "catalytic perfection", since it has a reaction rate controlled by diffusion. That is, the product formation takes place in a way as fast as the collision of the enzyme and substrate which limits the speed and is even spreading the product out of the active site of the enzyme.

Main bibliographic sources:
- Voet D, Voet JG, Biochemistry, Wiley
- Nelson DL, Cox MM, Lehninger - Principles of Biochemistry, WH Freeman Publishers

1 comment:

  1. Enzymes are proteins (some may be catalytic RNA) that have unique capacity of speeding up chemical reaction within cells. They accelerate the velocity of the reaction without being altered after the reaction. enzyme catalytic characteristics