Lecture 16: Regulation of protein function and enzymes Flashcards
Enzymes are:
proteins folded into complex shapes that allow substrate molecules to fit into the active site
Enzymes do not
change during reactions, nor do they change the other contents of the reaction. They simply facilitate the interaction of reactants and speed up the rate of the reaction.
Enzymes are the most:
selective and powerful catalysts known
hydrolases:
hydrolytic cleavage (including nucleases and proteases)
Nucleases :
break down nucleic acids by hydrolysing bonds between nucleotides
Proteases:
Break down proteins by hydrolysing bonds between amino acids
Syntheses:
Synthesise molecules by condensing two small molecules
Isomerases:
Rearrangement of bonds within a single molecule
Polymerases:
Polymerisation reactions such as synthesis of RNA and DNA
Kinases
Addition of phosphate to molecules (e.g. sugars, proteins)
Phophatases:
Removal of phosphate group
Oxide-Reductases:
One molecule is oxidised while another is reduced
ATPases:
Hydrolyse or synthesise ATP
Total number of enzyme in man is
~75,000
Many enzyme require:
cofactors, such as metal ions, NAD(P)(H), ATP, Vitamins etc
the function of over 60% of plant enzymes is
unknown
The ___ regulates the catalytic activity of enzymes
cell
regulation of enzyme protein abundance is by
transcription and translation
Direct fine-tuned control of protein activity and kinetics is by
metabolites and regulators (which could affect many enzymes)
Cell regulates enzymes for
specific post-translational modifications (specific covalent modifications affecting one single step)
Feedback inhibition in a metabolic pathway: bacterial pathway for tryptophan synthesis
- Trp can inhibit the activity of the first enzyme in the pathway (rapid response)
- or can repress expression of the genes for all the enzymes needed for the pathway (longer-term)
The first step of Trp biosynthesis in Arabidopsis is catalysed by
AS, anthranilate synthase, which is feedback inhibited by Trp.
Post-translation modifications of proteins -
PTMs
PTMs are..
rapid, highly regulated and highly specific. Over 300 types
example of PTMs
• addition of functional groups: simple (phosphorylation), or complex (glycosylation)
• structural changes: reduction (thiol/disulphides)
• changes to an amino acid: (e.g. Gln to Glu)
• addition of proteins or peptides: ubiquitination (small regulatory
protein that can signal degradation via the proteasome, alter cellular location, activity or protein interactions), SUMOylation (Small Ubiquitin‐like Modifier) that often increases the protein lifespan and stability.
PTMs: Proteins may undergo
a single type of modification or combinations of PTMs.
Some PTMs are
fixed for the life of the protein (cleavage of a signal peptide or glycosylation), while other changes are reversible, such as phosphorylation.
Post-translational modifications increase
protein diversity.
PTMs exponentially increase the complexity of the proteome relative to both the transcriptome and genome.
An efficient, cost‐effective mechanism for the diversification of the genome.
Moonlighting proteins
Many proteins have two or more different functions.
• Some crystallins, structural proteins in the lens of the vertebrate eye, are also enzymes, e.g. duck ε‐crystallin is lactate dehydrogenase, turtle τ‐ crystallin is enolase.
• Aconitase in man is both an enzyme of the Krebs cycle and involved in iron homeostasis.
Extended aerobic respiration requires
a regulated supply of carbon from glycogen and lipids
Glycogen granule has ____ glucose units and score protein ____.
~30,000
glycogenin
Glycogen is similar to _ in plants
amylopectin but is more branched
branching gives glycogen two advantages:
• more soluble.
• the exposure of more C4 (non‐
reducing) ends means that glycogen can be both synthesized and degraded more quickly than a single starch chain with the same number of residues.
Common PTM: Protein phosphorylation
twice that in mammals.
Kinase/Phosphotase cascades are common in
eukaryotes and act as amplifiers
Protein kinases make up ___ of the Arabidopsis genomes
~5.5%
twice that in mammals
Control of glycogen metabolism by protein phosphorylation: Glycogen breakdown increases
blood Glc → Glc 1‐P → Glc 6‐P. Glc 6‐P increases the rate of glycolysis in muscle.
Control of glycogen metabolism by protein phosphorylation: Insulin, glucagon, and epinephrine (adrenaline) are
hormones controlling glycogen metabolism in mammals. Glucagon and epinephrine stimulate glycogen degradation, opposing effect of insulin.
Glucagon is a
a peptide hormone, is secreted by the pancreas in response to low blood Glc. An elevated glucagon concentration occurs during fasting.
The adrenal glands release:
epinephrine in response to neural signals that trigger the fight‐or‐flight response (response to a sudden energy requirement).
Glucagon and epinephrine action is selective because
only liver cells have receptors. Glucagon and insulin act over longer periods to maintain a relatively constant concentration of blood Glc (~5 mM).
Diabetes: type 1
Type 1 diabetes develops when the pancreas is unable to produce any insulin (autoimmune disease).
diabetes: type 2
Type 2 diabetes develops when the pancreas can still make some insulin, but not enough, or when insulin that is produced does not work properly (insulin resistance).
Before discovery and purification of insulin
in the 1920’s diabetes was fatal.
Insulin was the first
peptide drug, protein sequenced, protein structure solved, hormone measured in blood, hormone gene cloned, first recombinant and first biotech drug.
Glycogen synthesis is regulated by protein phosphorylation: Glycogen synthase adds
UDP-Glc to a growing chain of glycogen. Two forms: glycogen synthase a is dephosphorylated and active. Glycogen synthase b is phosphorylated (P) by protein kinase A and is inactive.
Glycogen synthesis is regulated by protein phosphorylation: Protein kinase A is activated by
Protein kinase A (cyclic AMP‐dependent protein kinase A) is activated by a messenger, cyclic AMP (cAMP). The binding of epinephrine (adrenaline) or of glucagon to liver cell G‐protein linked receptors activates the adenylyl cyclase pathway which forms cAMP.
Kinase/ phosphatase cascades regulate glycogen metabolism:
Glycogen phosphorylase a is
active phosphorylated form of the enzyme that degrades glycogen. Glycogen phosphorylase b is the dephosphorylated inactive form.
Kinase/ phosphatase cascades regulate glycogen metabolism:
glycogen synthase and glycogen degradation have a
Reciprocal relationship of the glycogen synthase (inactive) and glycogen degradation enzymes (active).
Kinase/ phosphatase cascades regulate glycogen metabolism:
Phosphorylase kinase phosphorylates
Phosphorylase kinase phosphorylates glycogen phosphorylase. Phosphorylase kinase is activated by phosphorylation by protein kinase A. Thus, epinephrine and glucagon will stimulate glycogen degradation in addition to switching off glycogen synthesis.
The kinase cascade amplifies the
Signal.
Each successive element in the pathway, once activated, can act on many molecules of its substrate (next molecule in the cascade). This amplifies the signal and greatly increases the rate of glycogen mobilisation.
Insulin reverses
he effects of glucagon and epinephrine and stimulates glycogen synthesis in the liver
Insulin is synthesised
in the pancreas after feeding when blood glucose increases. High [Glc] lead to storage of carbohydrate as glycogen.
Effect of insulin is the opposite to
glucagon and epinephrine. Insulin binds to a cell surface receptor and triggers a pathway that leads to activation of protein phosphatase‐1. Dephosphorylation of three enzymes leads to inactivation of glycogen degradation and activation of glycogen synthesis.
