Molecular Modifications & Interactions Flashcards
Post-translational modification
refers to the covalent,
generally enzymatic, modification of proteins after
protein synthesis (translation)
➢ Post translational modifications can either modify an
existing functional group or introduce a new one
such as a phosphate group.
➢ Post-translational modifications can occur on
the side chains of the amino acids (R groups)
➢ Post-translational modifications can occur at
the protein’s C-terminal or N-terminal
Co-translational modification
refers to the covalent,
generally enzymatic, modification of proteins during
protein synthesis (translation).
Whats phosphorylation
➢ Phosphorylation is addition of phosphate group(s) to proteins.
(PO4)
➢ It is the most common protein modification.
➢ It is an important regulator of enzyme activity.
➢ Plays a key role in cellular signalling pathways.
Phosphate groups have other important functions in cells (non-protein functions)
- Phosphorylation is important in the metabolism of
carbohydrates - Phosphate group is a part of phospholipid structure
- Phosphate group is a component of DNA/RNA backbone
- Cyclic nucleotide monophosphates (cAMP) are important
signalling molecules
Common in bacterial signalling systems
Histidine: His~P
Aspartate: Asp~P
Important in enzyme activity
Common in eukaryotic signalling systems
Serine: Ser~P
Threonine: Thr~P
Tyrosine: Tyr~P
Enzymes in phosphorylation
➢ Enzymes that add phosphate group(s) are called KINASES.
➢ Enzymes that remove phosphate group(s) are called
PHOSPHATASES.
Chemistry of phosphorylation
➢ Phosphorylation introduces a
charged and hydrophilic group in the
side chain of amino acids.
➢ Phosphorylation can change a
protein’s structure by altering
interactions with nearby amino
acids.
Protein phosphorylation and enzyme activity
➢ Phosphorylation controls almost half of the enzymes
(activates/deactivates)
Example: Isocitrate dehydrogenase
Depending on carbon source this enzyme needs to be active/inactive.
Phosphorylation of a serine in the enzyme’s active site causes a loss of
activity.
Dephosphorylation restores activity.
Protein phosphorylation usually changes the function of the
target protein by
➢ Changing enzyme activity
➢ Changing its cellular location
➢ Altering its association with other proteins
Protein phosphorylation in cell signalling
➢ Protein phosphorylation plays a critical role in cell signalling
in response to extracellular stimulus and is very important
in biological regulation.
➢ Phosphorylation and de-phosphorylation commonly act as a
switch to turn on and off a signalling cascade.
Remember how phosphorylation of Rb by CDK inactivates Rb at G1/S
transition of the cell cycle, and its dephosphorylation by other
enzymes activates it.
Cell signalling and signal transduction pathways
➢ A signal transduction pathway is a sequence of molecular
events and chemical reactions that lead to a cell’s
response to a signal.
➢ Signal transduction pathways vary greatly in their details,
but every such pathway involves a signal, a receptor, and a
response.
➢ Protein phosphorylation/dephosphorylation play key roles
in signal transduction pathways.
Signal transduction in bacteria (phosphorelay)
➢ In bacteria, extracellular signals are transduced
into the cell predominantly by two-component
systems (TCSs).
➢ TCS consists of a sensor kinase – which is
membrane bound receptor protein, and a response
regulator protein.
➢ Sensor kinases are usually integral membrane
proteins that autophosphorylate from ATP at a
conserved histidine residue and then transfer the
phosphoryl group to a conserved aspartate in the
response regulator
Signal transduction in bacteria
The activated regulator then dimerises and activates the
transcription of target genes.
The Ras protein
➢ The name ‘Ras’ is an abbreviation of ‘Rat sarcoma’. Family
initially identified in rat cancers.
➢ Ras proteins are small membrane-bound
proteins.
➢ A short sequence at its C-terminus
contains a Cysteine residue which is
modified to have a fatty acid group
attached to it.
➢ Because of this fatty acid group, Ras is stably associated with the cell membrane.
➢ All Ras protein family members are
G proteins, i.e. they bind GTP and
belong to a class of proteins called
small GTPases.
➢ They are involved in transmitting
signals within cells (cellular signal
transduction).
➢ Because these signals result in cell
growth and division, overactive Ras
signalling can ultimately lead to
cancer.
➢ Ras mutations are detected in about
30% of human cancers.
N-Terminal acetylation
➢ About 85% of all human proteins and 68% in yeast are
acetylated at their N-terminus.
➢ Several proteins from Bacteria and Archaea are also
modified by N-terminal acetylation.
➢ N-terminal acetylation appears to have effects on:
* Protein stability
* Protein localisation (to Golgi)
* Protein synthesis (acetylation of ribosomal proteins)
Histone acetylation
➢ Proteins involved in supercoiling (storage of DNA).
➢ As DNA is negatively charged, the surface of histones
is very positively charged so the molecules can
interact.
➢ The primary sequence of human histone H1 protein is
40% lysine.
➢ Lysine residues toward the N-terminal tail of individual
histones are the target for acetylation.
➢ Histone acetylation (and deacetylation) has been shown to be
an important mechanisms in the regulation of gene
transcription.
➢ Histone acetylation activates transcription.
➢ Acetylation of histone protein tails in specific regions of
the brain appear to be crucial to the molecular basis of
addictions.
Protein methylation
➢ Protein methylation
typically takes place
on arginine or lysine
amino acid residues in
the protein sequence.
➢ Methyl groups are
added to terminal
amino groups by
methyltransferases
Example: Histone methylation
➢ Methylation of histones influences the way they interact
with DNA.
➢ The level of chromatin compaction depends heavily on
histone methylation (and other modifications).
➢ Methylated histones can either repress or activate
transcription depending on the site of methylation.
Example: Histone methylation mediated inactivation of transcription
IMPORTANT NOTE:
Histone methylation can inactivate or activate transcription. This example shows inactivation only.
Histone methylation and acetylation are examples of
epigenetic modifications
What are epigenetic changes?
Changes that affect gene expression without changing
the DNA sequence.
What are epigenetic modifications?
➢ Epigenetics is the study of heritable phenotype changes that
DO NOT involve alterations in the DNA sequence.
➢ Chromatin remodelling is often associated with epigenetic
modification.
➢ Remodeling is accomplished through two main mechanisms:
➢ Histone modifications: Post-translational modification of the
amino acids that make up histone proteins.
➢ DNA methylation: The addition of methyl groups to DNA
converting cytosine to 5-methylcytosine
DNA Methylation
➢ DNA methylation involves the addition of a methyl
group to a cytosine residue.
➢ From 1-5% of cytosines may be modified depending on
the organism.
➢ These C nucleotides (which are methylated) are typically
followed by a Guanine nucleotide (G). Therefore, these regions
are called CpG islands.
➢ CpG islands are abundant in promoter regions.
➢ DNA Methylation leads to inhibition of transcription.
DNA methylation location and effects
➢ Highly methylated areas tend to be less transcriptionally
active.
Is DNA methylation inherited?
➢ Patterns of DNA methylation can be inherited.
➢ When DNA is replicated only one strand contains methylcytosine.
➢ A maintenance methylase can catalyse the formation
methylcytosine in the new strand.
➢ Note: pattern can still be altered by demethylation
➢ DNA methylation is important in development. On
fertilisation many genes become demethylated and are
expressed.
➢ As cells develop genes whose products are no longer needed are “silenced” by methylation.
➢ Monozygotic (identical) twins start with same methylation
pattern but by age 50 patterns are quite different.
Indicates environment plays important role in epigenetic
modification.
PHOSPHORYLATION
AND GLUCOSE METABOLISM
➢ Glucose in blood plasma is maintained at a concentration of about 5 μM.
➢ But there is very little free glucose in cells.
➢ This is because free glucose is converted to glucose-6-
phosphate and trapped within the cell - BECAUSE the cell
membrane is negatively charged.
➢ The initial phosphorylation of glucose is required to increase
its reactivity.
➢ In glycolysis further phosphorylation takes place before
breakdown to pyruvate and net generation of 2 molecules of
ATP.
➢ Phosphorylation plays a key role in sugar metabolism.
