The following was written in response to a class biochemistry dicussion post.
PHK2 is an important enzyme in the regulation of blood sugar levels as it responds to the peptide hormones insulin and glucagon that are both released by the pancreas.
I’ve chosen the enzyme phosphofructokinase-2 (PHK2) and for 2 principle reasons: (1) I find this enzyme interesting because of how it differs from phosphofructokinase-1 (PFK1) in that it has a much lower affinity (greater K_m) for the same substrate glucose 6-phosphate (G6P) than PFK1 [1]. (2) I also find it interesting as it is part of at least 2 different pathways and I have found it difficult to follow not only how the enzyme itself works, yet how its product interacts with other metabolic pathways. I’ve been left with the question of “how does something with a low affinity for its substrate help to control blood-glucose levels in the body?” That is, why is something that is so bad at binding its substrate (locally) have such big effects on the body (globally)?
On balance, I hope that through this discussion I will be able to clarify these principles of action of PHK2 both locally and globally and I hope to be able to relate it into some of the symptoms of diabetes mellitus.
PHK2 is encoded by the phosphofructokinase bisphosphatase-1 (PHKB1) gene and comes in 3 classes [4]. The liver class (L-type), the muscle class (M-type) and the fibroblast class (f-type) [1]. Nevertheless we will focus on the liver as it is the major site of control of blood glucose levels in the body so that we might be able to generalize this discussion to diabetes mellitus, a group of metabolic disorder characterized by high blood-glucose level [5].
Morphologically speaking, PFK2 is an allosteric enzyme which has an active site for fructose 6-phosphate (F6P), its substrate [1]. F6P is converted (phosphorylated) to fructose 2,6-bisphosphate (F2,6-BP) when PHK2 is in the “off” position [1]. However, PFK2 can both phosphorylate and dephosphorylate F6P and F2,6-BP respectively. In particular, PFK2 is a bifunctional enzyme that catalyzes both the forward and reverse reaction depending on whether or not it has been turned “off” or “on” respectively [1]. When PHK2 is in the “on” position, the conversion (dephosphorylation) is from F 2,6-BP to F6P [1].
To clarify, turning the enzyme “on” literally means phosphorylating the enzyme while turning it “off” means the enzyme is de-phosphorylzed [4]. It follows that since PHK2 converts F6P to F 2,6-BP and yet, when it is “on” it does the reverse, we can see that the enzyme itself is inhibited by phosphorylation. Specifically, through the action of protein kinase A [1]. Additionally, it is inhibited by other cellular cues of high energy charge like: citrate and phosphophenol pyruvate (PEP). Conversely, it is excited by low ATP concentrations i.e. high AMP concentrations [1]. Furthermore, the metabolite of PFK2, F 2,6-BP, is a potent allosteric enhancer of PHK1 [1].
From the above complete overview abvove, we see that PHK2’s activity is affected by insulin and glucagon, and by ATP and AMP. It is known that its product F6P effects both the glycolysis and gluconeogensis [1]. It isn’t a wonder that given so many links to different metabolic pathways that it can be so confusing!
A summary:
With the above pathways, we know as the question, “what happens if PHK2 is inhibited?” I tried to find a real toxin or chemical that bound irreversible/covalently to the enzyme to kill it but I couldn’t find one. Nevertheless, inhibition in vivo is common and has mostly been addressed already.
Locally, inhibition of the PHK2 enzyme is the result of phosphorylation of protein kinase A which stabilizes the phosphatase activity on the enzyme as it is a bifunctional enzyme, i.e. it both phosphorylates and dephosphorylates, as a result less F 2,6-BP is produced and so the rate that glycolysis occurs at decreases [1].
This becomes more interesting however when the scope is increased to include blood-glucose levels as it is through PFK2 that glucagon and insulin function. Inhibition of PFK2 (turned “on” with phosphate) is associated with an increase in blood-glucagon while an activation of PFK2 (turned “off” without phosphate) is associated with an increase in blood-insulin levels [1]. This has important rammifications for diabetes mellitus of which there are different types.
Types I and II have symptoms that are similar to each other; however, they are caused by different mechanisms. Type I is caused as a result of little or no insulin production from birth, an autoimmune disorder etc. while type II is caused by a little insulin response from overexposure to insulin or other factors [5]. In both cases the symptoms are similar because both lead to hyperglycemia. Recall that PHK2 has a lower affinity for glucose and responds to both insulin and glucagon. In other words, it is one of the mediators for blood-glucose control as it acts more often under high blood-sugar conditions i.e. when the liver has access to a lot of glucose in a fed state meaning secretion of insulin [1]. Inhibition of PHK2 in this case would wreck the liver’s ability to know if there is a lot glucose present or not and as a consequence would not be able: to tell cells of the body take intake glucose, to turn off glycolysis, stop glycogen burn, create more glucose transporters etc [3]. The most intuitive symptom to explain from this train of cause-and-effects is fatigue after meals. After eating, blood sugar level increases but the cells are still burning energy and then burn glycogen which releases even more sugar into the blood i.e. a blood-glucose spike with no metabolic energy — fatigue after a meal [3].
There are other off-site affects of inapprorpiate PHK2 inhibition as well. For example, according to the hypothesis of DiNuzzo, astrocytes use a glycogenolytic pathway to produce glucose so that cerebral glucose can be saved for the neurons that need it and do not produce glycogen for energy storage [2]. In the context of their hypothesis, if PHK2 was inhibited astrocytes would compete for glucose as well and the exquiste glucose control in the brain would be damaged. I know this might be a bit of stretch. Feel free to read the paper and let me know what you think would happen to their hypothesis if PHK2 was inhibited!
Citations: