regulation of protein function Flashcards
why are proteins regulated?
to ensure:
maintenance of cellular homeostasis
responsiveness to the environment
efficiency
what are the major ways in which enzyme activity can be regulated?
- changes in substrate concentration
- binding of small effector molecules
- reversible covalent modifications
- binding of regulatory proteins
- proteolytic activation
- controlling the amount of enzyme present
how can controlling substrate concentration regulate protein function?
- affects rate
- particularly important when [S] is less than Km, when rate is linearly dependent on [S]
what are isoenzymes?
enzymes that catalyse the same reaction but have different amino acid sequences
what are the roles of isoenzymes?
- can have different metabolic uses in different organs
- can have different locations and metabolic roles in the same cell
- can have different roles at stages of development
- can have different responses to allosteric regulators
give an example of an isoenzyme
lactate dehydrogenase
- main types: H4 + M4
- H4 is found mainly in the heart, has low Km for lactate and favours lactate oxidation
- M4 is found in skeletal muscle, has low Km for pyruvate and favours pyruvate reduction
what is allosteric regulation?
regulation mediated by interactions of a modulator at a regulatory site away from the active or binding site. binding of a modulator causes a conformational change
what is a homotropic modulator?
an allosteric modulator which is the same as the substrate
what is a heterotropic modulator?
an allosteric modulator which is different from the substrate
do allosterically modulated enzymes obey Michaelis-Menten kinetics?
no
give an example of allosteric regulation
co-operative oxygen binding in haemoglobin
haemoglobin undergoes a structural change on binding Hb, oxygen binding promotes stabilisation of high affinity R state
what do allosteric activators do?
increase the proportion of enzyme in the R state
what do allosteric inhibitors do?
increase the proportion of enzyme in the T state
when are covalent modifications made?
post-translationally
how do protein kinases cause covalent modification?
protein kinases transfer their terminal phosphate from ATP to the OH group of Ser/Tyr/Thre
how do protein phosphatases cause covalent modification?
catalyse the hydrolytic removal of phosphoryl groups from proteins
why is phosphorylation an effective covalent modification for regulation of protein?
- large free energy permits changes in conformation of phosphorylated protein
- addition of 2 negative charges disrupts existing electrostatic interactions
- phosphoryl group can form 3 new H bonds, allowing specific interactions with hydrogen bond donors
- permits amplification cascades
explain how phosphorylation by kinases results in signal amplification?
activation of enzymes by other enzymes increases number of affected molecules - allowing signal amplification of several orders of magnitude in a few ms
how is PKA an example of phosphorylation regulation?
in the absence of cAMP a pseudosubstrate region blocks activity of PKA
PKA consists of regulatory subunit and catalytic subunit
when cAMP binds to R subunit, it dissociates, releasing the C subunit which is catalytically active
what is the concerted model for allostery?
one substrate molecule causes complete change in R state
what is the sequential model for allostery?
substrate binding to one subunit will only affect adjacent subunits
why is CTP an ATCase inhibitor?
CTP favours the R state, stabilising the inactive state
what is the difference in response for allosteric enzymes vs those which obey Michaelis-Menten kinetics?
allosteric enzymes can respond to small changes in [S] compared to those which obey michaelis-menten
how do adaptor proteins spatially restrict signals in the cell?
a-kinase anchoring proteins have different domains specific for PKA, PP2A, the target protein and specific regions of the cell membrane
describe the structure of calmodulin
calcium is bound between two a-helices and a short loop, it binds via 7 oxygen atoms. there are 4 EF hand motifs to provide 4 Ca2+ binding sites.
how is calmodulin regulated by calcium?
calcium binding to calmodulin causes a conformational change in the protein, and binds exposed a-helices on the target protein
how does calcium regulate protein phosphatase calcineurin?
in the inactive state, an auto inhibitory domain blocks the active site, calcium binding triggers a change in conformation that unblocks the active site
how can proteins be regulated by partial proteolysis?
precursors (pro proteins or zymogens) can be synthesised and removed in a highly specific, irreversible process to release the active protein
how are active zymogens switched off?
tight binding of inhibitors or degradation
how can the amount of protein present in the cell be regulated in the long term?
change in the rate of protein synthesis, enzyme repression
how does the ubiquitin-proteasome pathway affect the regulation of the amount of protein in the cell?
changes the rate of protein degradation
how can the catalytic activity of enzymes be controlled?
feedback regulation
