Metabolism 1/2 (handout p. 21-45) Flashcards
Glucose, galactose and fructose (hexoses) play a central role in energy homeostasis in high
mammals. However, these molecules are unable to diffuse passively across cellular membranes, and
require transporter proteins for entry into and exit from cells. Describe the two distinct groups of transporters that
have been identified.
1.Hexose transporters down a concentration gradient (GLUT1, GLUT2, GLUT3, GLUT4
and GLUT5).
- Hexose transporters against a concentration gradient using energy provided by an
electrochemical gradient of sodium, which is co-transported with the hexose (SGLT1,
SGLT2).
Describe the affinities of the transporters for glucose, galactose and fructose. Which have a high affinity for glucose (what value indicates this?) Which has a low affinity for glucose?
Class I (GLUTs 1-4) are glucose transporters. Class II (GLUTs 5, 7, 9 and 11) are fructose transporters. Class III (GLUTs 6, 8, 10 and 12) are structurally atypical members of the GLUT family, which are poorly defined at present.
Each of the transporters has different affinities for glucose and the other hexoses, which largely
dictates their function. GLUT1, 3 and 4 have a high affinity for glucose (Km value = 2-5 mM, an
indicator of the affinity of the transporter protein for glucose molecules; a low Km value suggests a
high affinity), which indicates that they are functioning at maximal rate under physiologic
concentrations of glucose (approx. 5 mM). In contrast, GLUT2 has a low affinity for glucose (Km approx. 15
mM), which allows it to change transport rate in proportion to the increasing glucose concentrations
that occur after ingestion of a carbohydrate-rich meal.
How are GLUT transporters componsed. Describe the transport of glucose (saturable? direction? what drives it?).
In a resting post absorptive state, how is glucose metabolized?
GLUTs are composed of 12 membrane-spanning helices with an intracellular loop connecting the
6th and 7th helices. The facilitative transport of glucose is saturable, stereoselective and
bidirectional. Uptake of glucose by these transporters is along concentration gradient as intracellular glucose is actively metabolized by hexokinase and glucokinase. In resting post absorptive state about 70% glucose is metabolized in an insulin independent manner. This insulin-dependent and mostly independent mechanism is impaired in Type 2 Diabetes mellitus and in normoglycaemic subjects with family history of diabetes (glucose resistance).
Describe SGLT1.
What does it transport? What does it not transport? Where is it expressed?
Sodium glucose transporter 1; Co-transports one molecule of glucose or galactose along with two sodium ions. Does not transport fructose; Expressed in intestinal
mucosa, kidney tubules. It is insulin independent.
Describe SGLT2.
What does it transport? What does it not transport? Where is it expressed?
Sodium glucose transporter 2; Co-transports glucose and sodium ions. Does not
transport fructose or galactose; Expressed in kidney tubules.
Describe GLUT1.
What type of transporter? Where is it expressed?
Insulin independent plasma membrane transporter in RBCs (erythrocytes), brian and
endothelial cells. It is also widely distributed in fetal tissues
Describe GLUT2.
What type of transporter? Where is it expressed?
Insulin independent low affinity, high capacity transporter in the liver, also found in
the intestines and kidney (Bidirectional allowing glucose to flow in 2 directions).
Also, serves as a “glucose sensor” in pancreatic beta cells. GLUT 2 transports glucose
out of intestine, into the bloodstream, and into the liver. It is important to note that the
liver sees really high concentrations of glucose.
Describe GLUT3, GLUT4, GLUT5.
3- Insulin independent transporter in brain
4- INSULIN DEPENDENT transporter in MUSCLE, HEART and ADIPOCYTES.
Higher affinity for glucose. It gets glucose after we have eaten. This transporter is
not active during the fasting state.
5- Fructose transporter in skeletal muscle, adipose tissue, brian, sperm and erythrocytes
Which transporter is insulin independent low affinity, high capacity transporter in the liver, intestines and kidney?
GLUT2
The following describes which transporter:
INSULIN DEPENDENT transporter in MUSCLE, HEART and ADIPOCYTES.
Higher affinity for glucose.
GLUT4
Compare GLUT1 and GLUT 3 against GLUT2 and GLUT4 on a graph that compares glucose concentration to transport rate (% of Vmax).
Slide 32 or p 23
Glycolysis is regulated in three steps. Does it involve equilibrium or non-equilibrium
reactions? The enzymes that catalyze these steps are regulated in three
ways, describe.
Draw a chart w the steps.
non-equilibrium
allosterically, by covalent modifications, an regulation of the
amounts/synthesis of the enzymes.
flow chart p 24
Give an overview of glycolysis.
Partial breakdown of glucose into what? Describe the outputs.
What is produced?
Glycolysis is defined as a partial breakdown of glucose into 2 molecules of pyruvic acid, 4
protons and 4 electrons (accepted by 2NAD+ to form 2NADH + 2H+), and 2 net ATP (from
substrate-level phosphorylation). Note: it is important to know that the reaction actually goes
through twice resulting in the production of 4 ATP.
Where does glycolysis take place?
It produces energy in what form?
Glycolysis takes place in the cytoplasm, and, therefore, mitochondria are not required. It is
active in all cell types.
Glycolysis produces energy in the form of ATP and NADH.
Glycolysis is not isolated from other metabolic pathways.
Other molecules besides glucose can enter at a few points along the glycolytic pathway. Describe.
For example, the product of glycogen breakdown, glucose-6-phosphate, can enter the
glycolytic pathway at the second step. Glyceraldehyde-3-phosphate, which is produced by
photosynthesis, is also a glycolytic intermediate, so it can be directed from this anabolic
pathway into glycolysis when energy is needed.
Intermediates can be drawn out of the glycolytic pathway when energy levels are high, for use in biosynthetic pathways. What is an example?
during active energy
production pyruvate, the product of glycolysis, enters the citric acid cycle, but when energy
is not needed pyruvate serves as a substrate in amino acid synthesis.
At what point/step in glycolysis is glucose trapped in the cells?
Once glucose is phosphorylated into glucose 6-phosphate (in the first step of glycolysis), it is
trapped in the cells.
What enzyme mediates the most highly regulated part of glycolysis?
Aldolase A mediates the most highly regulated part of glycolysis (the splitting stage shown
below).
Describe the role of the following glycolytic enzymes.
Which use ATP and which generate ATP?
What does deficiency of glycolytic enzymes lead to?
- Hexokinase/glucokinase – catalyzes the first reaction in glycolysis, which is the
phosphorylation of glucose into glucose 6-phosphate. - PFK-1 – is highly regulated and is important in locking in the glycolytic pathway.
- Pyruvate Kinase
All the three enzymes mentioned above catalyze irreversible steps. It is important to note that the
first two enzymes (Hexokinase/glucokinase and PFK-1) utilize ATP, and the last two (Pyruvate
Kinase and Phosphoglycerate kinase) generate ATP.
The deficiency of glycolytic enzymes can lead to hemolytic anemia. Red blood cells burst, lose
hemoglobin, and you become anemic.
During what step does substrate level phosphorylation occur?
Substrate level phosphorylation occurs in the step when glyceraldehyde 3-phosphate is converted to 1,3-Bisphosphoglycerate and then 3-Bisphosphoglycerate by the enzyme Phosphoglycerate kinase.
Draw/explain the 3 stages of glycolysis.
P 26
What is allosteric regulation?
Reversible? Speed?
What effect might a buildup of the product have?
activation or inhibition of enzyme activity. The molecule or ligand
interacts with the enzyme at the active site and can either speed up the reaction or slow it
down. Allosteric regulation is reversible and is usually quick and transient. A buildup of
product can inhibit enzyme activity.
p27
Describe covalent modification and discuss the prime example.
Phosphorylation is initiated by a hormone and occurs when a phosphate group is
bonded with a hydroxyl group on the enzyme, adding a negative charge to the enzyme. The large negative charge initiates a conformational change, which then changes the activity of the enzyme. An example is the activation of protein kinase A (see picture below), which
phosphorylates syrine or threonine hydroxyl groups.
The phosphate is stuck on the hydroxyl group until it is removed by a phosphatase.
For example, insulin activates phosphatases to inhibit epinephrine and glucagon activity.
p 27, slide 39
For the following stimuli, what is the kinase?
hormone
neurotransmitter
cytokine
growth factor
See table on slide 40/ p 28
How are enyzmes synthesized?
Slide 42, p 28
This usually takes longer (can take many hours). A long-time fasting state or high consumption
of carbohydrates will initiate change in the enzyme activity.
Where is hexokinase present? Km?
How does it interact with glucose?
Hexokinase (a.k.a. Hexokinase Isozymes I, II and III):
- present in all cell types
- allosterically inhibited by its product, glucose
6-phosphate (e.g. feedback inhibition)
- constitutive enzyme, non-inducible, constant amount
low Km for glucose approx. 0.1 mM; saturated at low
glucose concentrations
- present at constant levels all the time, whether or not they
are activated.
- Hexokinase cannot handle very high levels of glucose.
Describe glucokinase. Where is it present?
What effect does fructose-6-phosphate have?
Km for glucose?
Glucokinase (a.k.a. Hexokinase Isozyme IV):
- present in liver and pancreas
- Translocation between nucleus (inactive) and cytosol (active)
- fructose 6-phosphate (downstream product) decreases activity by
promoting translocation to nucleus
- glucose increases activity by promoting
translocation to cytosol
- inducible enzyme; enzyme synthesis
induced by insulin — increased amount of
enzyme due to insulin
-Insulin increases expression of the gene.
- high Km for glucose ~ 7-10 mM;
not saturated at normal physiological
glucose concentration
-Glucokinase can handle large concentrations
of glucose in the liver.
Note: Blood glucose levels can get up to 5nM
