Topic 10: Protein function Flashcards
List the major regulatory mechanisms that control enzyme activity, and state examples
- Different enzyme forms-isoenzymes, enzymes that catalyse same rxn but have different AA sequence, diff activity and regulatory properties. Synthesized from different genes or differentially spliced from same gene.
- Change in enzyme configuration- allosteric regulation, usually muti-subunit, can exist in 2 different forms T( low affinity) or R state. Activators or inhibitors( increase proportionof enzyme in T state, ROR decreases). Activators: AMP, fructose 2,6 biphosphate. Inhibitors: ATP, citrate, H+.
- Reversible covalent modification- phosphorylation, protein kinase( transfer terminal phosphate from ATP to OH group of Ser, Thr, Tyr) OR protein phosphatases( reverse effects of kinases by catalysing hydrolytic removal of phosphoryl groups from proteins.
- Proteolytic activation
- Controlling the amount of enzyme present- gene expression
Discuss the allosteric properties of a key regualtory enzyme
R and T states: R( high affinity), activator, curve shifts to left.
Allosteric effectors
Examples:
activators: AMP, fructose 2,6-biphosphate
inhibitors: citrate, H+, ATP
Discuss the concept of enzyme cascades and use of protein kinases and phosphatases to regulate activity
Why cascades are impt: when enzymes activate enzymes, the number of affected molecules increases geometrically in an enzyme cascade.
What is kinase/phosphatase: protein kinase removes terminal phosphate from ATP and moves it to OH group of Ser, Thr, Tyr. Phosphatase reverses this rxn.
Why does phosphorylation have an effect: the free energy of phophorylation is large, adds 2 negative charges, a phosphoryl group can make H bonds which can lead to other interactions being formed. Rate of phosphorylation and dephosphorlation can be adjusted. This process links energy status of cell to matabolism through ATP, and allows for amplification effects
Define the term zymogen with examples
An inactive precursor molecule, aka proenzyme in the proteolytic activation process. Involves the breaking of a peptide bond, irreversible, important when process need to be tightly controlled.
eg. Blood clotting, digestive enymes, apoptosis( programmed cell death)
eg: pancreatic zymogens whose activation is controlled by trypsin set off a chain of reactions which activates many other enzymes like lipase, elastase, carboxi-peptidase
Explain how activation of clotting cascade leads to the formation of fibrin
- Intrinsic pathway: damaged endothelial lining of blood cells promotes binding of factor 12. ie blood vessels are damaged. This leads to activation of factor 10, thrombin activation and formation of fibrin clot
- Extrinsic pathway: trauma releases tissue factor 3, factor 10 activated, thrombin activated, formation of fibrin clot
Discuss mechanisms involved in the regulation of clot formation
- Prothrombin to thrombin
2 kringle domains in help keep prothrombin in inactive form. Gla domains target it to appropriate sites for activation. - Fibrinogen to fibrin
Fibrinogen is converted by thrombin into fibrin: composed of 3 polypeptide chains and have 2 globular heads. Fibrinopeptides prevent fibrinogen molecules coming together. Thrombin cuts into fibrinopeptides to produce fibrin. Fibrin monomers assemble by non-covalent interactions to form soft clot. Cross linking of soft clots by covalent bonds catalysed by transgulataminase. - Sustained activation of pathway
Factors 5 and 8 stimulate activity of other ezymes throughout pathway. Thrombin positive feedback on factors 5 etc
Explain the physiological roles of myoglobin and haemoglobin
Contrast the oxygen binding properties of myoglobin and haemoglobin, and explain why haemoglobin is most suited to its role as O2 transporter
Myoglobin: constant affinity for oxygen, hyperbolic curve
Haem: affinity for O2 increases as partial pressure of O2 increases. Haem undergoes conformational change on binding with oxygen, makes it easier for more 02 to bind with haem. Sigmoidal curve
Describe major structural differences between oxygenated and deoxygenated haemoglobin, and the molecular basis of cooperativity
Cooperativity: enhances O2 transport by haem. Binding with O2 promotes transition from low affinity T state to high affinity R state. Sigmoidal properties of Hb allows greater O2 transport compared to comparable proteins entirely in the high affinity R state or low affinity T.
Describe the effects of CO2, H+, 2,3-biphosphoglycerate and carbon monoxide on the binding of O2 to haem, and the physiological significance of these effects
BPG(-ve): lowers afinity of Hb for oxygen, stabilises T state as it interacts with +ve residues on each beta haem subunit. Shifts curve to right
CO2 and H+( Bohr effect): Binding of CO2 and H+ lowers affinity of Hb for O2. Stabilises T state, allows delivery of O2 to metabolically active tissues that produce CO2 and H+ like muscles. Shifts curve to right
CO: binds to haem 250x more readily than O2. Blocks O2 from binding with haem, stabilises in R state, prevents dissociation at tissues, O2 deprivation. Shifts curve to left.
What is the significance of mutations in globin genes?
Gives rise to diseases such as sickle cell anaemia- wrong amino acid in chain. Cells formed are more prone to lyse- leads to anaemia. They are also more rigid, and will be unable to fold and move through smaller capillaries.
Outline key features needed for protein secretion
Describe differences between constitutive and regulated secretion
In regulated secretion, secretory materials are stably accumulated in secretory vesicles as storgage sites. In constitutive secretion, secretory materials are continuously released
Outline structure of collagen
triple helical arrangement of collagen chains. Contain Gly-X-Y repeating sequence. Hydrogen bonds stabilise interactions between chains. Collagen fibrils formed from covalently cross-linked collagen molecules.
Outline processing of collagen
