Several models have been developed for relating changes in dissociation constants to changes in the tertiary and quaternary structures of oligomeric proteins. One model suggests that the protein's subunits can exist in either of two distinct conformations, R and T. At equilibrium, there are few R conformation molecules: 10 000 T to 1 R and it is an important feature of the enzyme that this ratio does not change. The substrate is assumed to bind more tightly to the R form than to the T form, which means that binding of the substrate favors the transition from the T conformation to R. The conformational transitions of the individual subunits are assumed to be tightly linked, so that if one subunit flips from T to R the others must do the same. The binding of the first molecule of substrate thus promotes the binding of the second and if substrate is added continuously, all of the enzyme will be in the R form and act on the substrate. Because the concerted transition of all of the subunits from T to R or back, preserves the overall symmetry of the protein, this model is called the symmetry model. The model further predicts that allosteric activating enzymes make the R conformation even more reactive with the substrate while allosteric inhibitors react with the T conformation so that most of the enzyme is held back in the T shape.Experiment Evaluating Non-Symmetry Model EnzymesExperiments were performed with enzyme conformers that did not obey the symmetry model. The data is summarized in Figure 1.Equilibrium distribution of two conformers at different temperatures given the free energy of their Figure 1:interconversion. (modified from Mr.Holmium).Allosteric enzymes differ from other enzymes in that they:
Answer(s): B
You must be familiar with how enzyme function is regulated to answer this question. An allosteric enzyme has a site other than the one for the substrate at which a molecule (not the substrate) that directs the function of the enzyme can bind. The above illustrates the allosteric regulation of an enzyme with a positive effector (on the left) and a negative effector (on the right).
Several models have been developed for relating changes in dissociation constants to changes in the tertiary and quaternary structures of oligomeric proteins. One model suggests that the protein's subunits can exist in either of two distinct conformations, R and T. At equilibrium, there are few R conformation molecules: 10 000 T to 1 R and it is an important feature of the enzyme that this ratio does not change. The substrate is assumed to bind more tightly to the R form than to the T form, which means that binding of the substrate favors the transition from the T conformation to R.The conformational transitions of the individual subunits are assumed to be tightly linked, so that if one subunit flips from T to R the others must do the same. The binding of the first molecule of substrate thus promotes the binding of the second and if substrate is added continuously, all of the enzyme will be in the R form and act on the substrate. Because the concerted transition of all of the subunits from T to R or back, preserves the overall symmetry of the protein, this model is called the symmetry model. The model further predicts that allosteric activating enzymes make the R conformation even more reactive with the substrate while allosteric inhibitors react with the T conformation so that most of the enzyme is held back in the T shape.Experiment Evaluating Non-Symmetry Model EnzymesExperiments were performed with enzyme conformers that did not obey the symmetry model. The data is summarized in Figure 1. Equilibrium distribution of two conformers at different temperatures given the free energy of their Figure 1:interconversion. (modified from Mr.Holmium).All of the following statements are consistent with Figure 1 EXCEPT:
Answer(s): D
Conformational isomers exist in a dynamic equilibrium, where the relative free energies of isomers determine the population of each isomer and the energy barrier of rotation determines the rate of interconversion between isomers.Answer choice A: The graphs shows that for exergonic reactions (this means G < 0), the equilibrium constant is always greater than 1 (meaning that the forward reaction is favored which implies that the products are Kmore stable). Recall that the equilibrium constant K is defined as the product of the product concentrations divided by the product of reactant concentrations.Answer choice B: divide both sides by -RT then raise both sides to the power of e. Remember from your rules of logarithms than e to the power of ln x = x. Also recall that ln is log to the base e.Answer choice C: From the graph, at a free energy difference of 0 kcal/mol (= equilibrium), this also gives an equilibrium constant of 1.Answer choice D: A negative difference in free energy means that a conformer interconverts to a thermodynamically more stable conformation, thus the equilibrium constant will always be greater than 1.However, notice that K decreases with increasing temperature meaning that there is more, relatively, of the less stable conformer (in other words, energy from the higher temperature is able to override the energy barrier to conversion to the less stable conformer). Thus the statement in answer choice D is incorrect which makes answer choice D the correct answer!Going a bit further: notice that a positive difference in free energy means the conformer already is the more stable one, so the interconversion is an unfavorable equilibrium (K < 1). Even for highly unfavorable changes (large positive G), the equilibrium constant between two conformers can be increased by increasing temperature, meaning the amount of the less stable conformer present at equilibrium does increase slightly (see graph).Note that: An endergonic process is accompanied by or requires the absorption of energy, the products being of greater free energy than the reactants. An exergonic process is the opposite and thus accompanied by the release of energy. Incidentally, the enzyme subunits described in the passage exist in one of two conformations which stands for 'tensed' (T) or 'relaxed' (R), and as described relaxed subunits bind substrate more readily than those in the tense state. At any rate, T/R was not to be considered in this question.
The polymerase chain reaction (PCR) is a powerful biological tool that allows the rapid amplification of any fragment of DNA without purification. In PCR, DNA primers are made to flank the specific DNA sequence to be amplified. These primers are then extended to the end of the DNA molecule with the use of a heat-resistant DNA polymerase. The newly synthesized DNA strand is then used as the template to undergo another round of replication.The 1st step in PCR is the melting of the target DNA into 2 single strands by heating the reaction mixture to approximately 94° C, and then rapidly cooling the mixture to allow annealing of the DNA primers to their specific locations. Once the primer has annealed, the temperature is elevated to 72° C to allow optimal activity of the DNA polymerase. The polymerase will continue to add nucleotides until the entire complimentary strand of the template is completed at which point the cycle is repeated (Figure 1)Figure 1One of the uses of PCR is sex determination, which requires amplification of intron 1 of the amelogenin gene. This gene found on the X-Y homologous chromosomes has a 184 base pair deletion on the Y homologue. Therefore, by amplifying intron 1 females can be distinguished from males by the fact that males will have 2 different sizes of the amplified DNA while females will only have 1 unique fragment size.The polymerase chain reaction most likely resembles which of the following cellular process?
Answer(s): C
As described in the passage, the polymerase chain reaction utilizes DNA primers that anneal to the DNA template. DNA polymerase then extends the DNA primers in an effort to replicate the DNA strand. These steps are identical to those of DNA replication in a cell. However, in a cell, the cycle occurs once per cellular division.
The polymerase chain reaction (PCR) is a powerful biological tool that allows the rapid amplification of any fragment of DNA without purification. In PCR, DNA primers are made to flank the specific DNA sequence to be amplified. These primers are then extended to the end of the DNA molecule with the use of a heat-resistant DNA polymerase. The newly synthesized DNA strand is then used as the template to undergo another round of replication.The 1st step in PCR is the melting of the target DNA into 2 single strands by heating the reaction mixture to approximately 94° C, and then rapidly cooling the mixture to allow annealing of the DNA primers to their specific locations. Once the primer has annealed, the temperature is elevated to 72° C to allow optimal activity of the DNA polymerase. The polymerase will continue to add nucleotides until the entire complimentary strand of the template is completed at which point the cycle is repeated (Figure 1)Figure 1One of the uses of PCR is sex determination, which requires amplification of intron 1 of the amelogenin gene. This gene found on the X-Y homologous chromosomes has a 184 base pair deletion on the Y homologue. Therefore, by amplifying intron 1 females can be distinguished from males by the fact that males will have 2 different sizes of the amplified DNA while females will only have 1 unique fragment size.Why is a heat resistant DNA polymerase required for successive replication in the polymerase chain reaction, rather than simply a human DNA polymerase?
Answer(s): A
Since mammalian cells function at 37° C, this is also the optimal temperature for enzyme activity. The temperatures used in PCR, which are well above 70° C, would easily denature human DNA polymerase which is why a heat resistant DNA polymerase is required. Using human DNA polymerase would require new enzyme after each cycle, and therefore would not be very efficient.
The polymerase chain reaction (PCR) is a powerful biological tool that allows the rapid amplification of any fragment of DNA without purification. In PCR, DNA primers are made to flank the specific DNA sequence to be amplified. These primers are then extended to the end of the DNA molecule with the use of a heat-resistant DNA polymerase. The newly synthesized DNA strand is then used as the template to undergo another round of replication.The 1st step in PCR is the melting of the target DNA into 2 single strands by heating the reaction mixture to approximately 94° C, and then rapidly cooling the mixture to allow annealing of the DNA primers to their specific locations. Once the primer has annealed, the temperature is elevated to 72° C to allow optimal activity of the DNA polymerase. The polymerase will continue to add nucleotides until the entire complimentary strand of the template is completed at which point the cycle is repeated (Figure 1)Figure 1One of the uses of PCR is sex determination, which requires amplification of intron 1 of the amelogenin gene. This gene found on the X-Y homologous chromosomes has a 184 base pair deletion on the Y homologue. Therefore, by amplifying intron 1 females can be distinguished from males by the fact that males will have 2 different sizes of the amplified DNA while females will only have 1 unique fragment size. Which of the following statements could be used to correctly describe the overall polymerase chain reaction?
This question requires knowledge of the definition of anabolism and catabolism. A catabolic reaction involves the breakdown of macromolecules, whereas an anabolic reaction involves the synthesis of macromolecules from individual building blocks. PCR entails the synthesis (amplification) of a new DNA strand using a DNA template and free nucleotides, therefore, it is an anabolic reaction that synthesizes new DNA strands.
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