T11.2 - Regulatory Mechanisms Flashcards
what are regulatory mechanisms
the way in which we control the activity of enzymes and proteins
often involve…
- direct regulation of enzyme
- gene expression (not really covered here)
what are some examples of short term regulation
- isoenzymes: different enzyme forms
- allosteric regulation: change in enzyme conformation
- phosphorylation: reversible covalent modification
- proteolytic activation
- controlling the amount of enzyme present by gene expression (not covered much here)
what are isoenzymes
different forms of the same enzyme
* they catalyse the same reaction but have different amino acid sequence
* different activity
* different regulatory activity (eg Km)
* therefore can adapt to needs of different tissues
* synthesised from different genes or differently spliced from the same gene
eg hexokinase and glucokinase (in the liver) - both can catalyse the phosphorylation of glucose, but hexokinase has a much lower Km value so can reach Vmax very quickly
allosteric regulation
change in enzyme conformation
* allosteric regulator binds at a site away from the active site
* results in conformational change that alters the properties of the protein
* usually multi-subnit
* idea is that enzyme exists in two different forms…
T state - low affinity
R state - high affinity
- activators increase the proportion of the enzyme in the R state → shift curve to left → more molecule in higher affinity form → increases activity
- inhibitors increase proportion of enzyme in T state → curve shifts to right → lower activity
note: allosteric regulation doesn’t obey Michaelis-Menten kinetics, as sigmoidal curve here is different shape to what we’d see in M-M kinetics
allosteric = means ‘action at a distance’
example of allosteric regulation: phosphofructokinase-1
fructose 6 phosphate → fructose 1,6 bisphosphate
activators
* AMP
* fructose 2,6-bisphosphate
* these shift the curve to left, increasing activity of enzyme
inhibitors
* citrate
* ATP
* H+
* these shift curve to right, decreasing activity of enzyme
what is reversible covalent modification - phosphorylation
the addition of a phosphate group
* derived from ATP molecule
* added onto specific amino acid residues (those with hydroxyl group)
* eg ATP added to -OH of Ser, Thr, Tyr
* this is done by protein kinases
* there are many different types of PKs, each with different specificity (look for different AA sequences)
* this is a reversible process
* can take phosphates off by protein phosphatases
* PPs reverse effects of kinases by catalysing the hydrolytic removal of phosphryl groups from proteins
what is reversible covalent modification - phosphorylation
the addition of a phosphate group
* derived from ATP molecule
* added onto specific amino acid residues (those with hydroxyl group)
* eg ATP added to -OH of Ser, Thr, Tyr
* this is done by protein kinases
* there are many different types of PKs, each with different specificity (look for different AA sequences)
* this is a reversible process
* can take phosphates off by protein phosphatases
* PPs reverse effects of kinases by catalysing the hydrolytic removal of phosphryl groups from proteins
allows for amplification effects (explained on another card)
why is protein phosphorylation so effective?
- free energy of phosphorylation is large involves hydrolysis of ATP → large neg change in free energy → able to have change in protein conformation
- adds 2 neg charges can enhance/disrupt interactions within molecule itself or with other molecules
- a phosphoryl group can make H-bonds which allows for interactions between enzyme subunits, within same subunit or with other regulatory molecules
- rate of phosphorylation/dephosphorylation can be adjusted by changing relative activity of kinases and phosphatases to fine tune to what cell needs
- links energy status of cell to metabolism by ATP energy status of cells gets linked to phosphorylation of many key enzymes and proteins
- allows for amplification effects where one activated enzyme can activate several more, which activates several more etc… cascade like mechanism (explained on next card)
amplification effects, eg phosphorylation
when enzymes activate enzymes, the number of affected moleules increasees in an enzyme cascade
- eg signal outside cell binds to receptor
- this activates enzyme 1
- enzyme 1 phosphorylates several of enzyme 2
- each enzyme 2 goes on to phosphorylate several other enzyme 3s
- amplification of signal
- many enzymes activated very quickly
- leads to cascade effects
what is proteolytic activation
- precursors are completely inactive when they are in zymogen/proenzyme form
- they have different AA sequence to the final activated form
- proteolytic activation involves breaking of peptide bond
- this takes off the prosegment (part of original zymogen/proenzyme)
- this activates, converts zymogen to active form
- irreversible, cannot put prosegment back on
- therefore need regulation of activated molecule too
- important when processes need to be tightly controlled
eg blood clotting, digestive enzymes and apoptosis
example is pancreatic zymogens (next card)
proteolytic activation example - pancreatic zymogens
- pancreas is major source of digestive enzymes
- only want to activate digestive enzymes when they are outside of the cell
- trypsinogen (trypsin inactive form) is secreted into pancreatic duct
- enteropeptidase cuts off small part of trypsinogen to form active trypsin
- trypsin then goes onto stimulate activation of other zymogens to activate other digestive enzymes
what state of enzymes are high and low affinity
T state
low affinity - lower activity at any given substrate concentration
R state
high affinity - greater activity
what do allosteric inhibitors/activators stabilise
in terms of T and R state, shift curve to…
activators
- increase proportion of enzyme in R state by stabilising R state
- shift curve to left
- there is more molecule in higher affinity form
- therefore higher activity
inhibitors
- increase proportion of enzyme in T state by stabilising T state
- shift curve to right
- more molecule in lower affinity form
- therefore lower activity
