Prevent theft first, and replace it later.

1.3: The isoelectric point of tyr is (2.29.1)/2=5.65, and the isoelectric point of cys is (2.08.4)/2=5.2

1.4: If anion exchange chromatography is selected, ASP, Lys and Ala are loaded on the column under a low salt buffer solution of pH6, then the negatively charged ASP will be adsorbed on the resin, while the positively charged Lys and the net charge of zero ALA will appear directly in the effluent, and ASP will be obtained in the subsequent elution of the high-salt buffer solution, so that the separation of ASP from Lys and Ala will be realized. If LYS and ALA need to be separated, the collected direct effluent can be subjected to cation exchange chromatography.

2.5: (4) It is the most stable, because the N-terminal and C-terminal of this A-helix have negatively charged and positively charged side-chain groups respectively, which can effectively neutralize the positive and negative charges at the N-terminal and C-terminal brought about by the dipole moment of the helix; (3) It is the most unstable, because the A-helix N-terminal and C-terminal have positively charged and negatively charged side-chain groups, which not only cannot neutralize the positive and negative charges at the N-terminal and C-terminal brought about by the helical dipole moment, but instead increases the charge at both ends.

2.6: The amino acid residues that appear at the two-phase interface should not be the amino acid residues that destroy the helix, and secondly, the side chains of this amino acid are relatively large and have amphiphilic properties - one part is hydrophilic and the other part is hydrophobic. Apparently TYR and LYS have such a nature.

2.11: The first should be an amino acid that is conducive to the formation of an a-helix; The second side chain should be an amino acid containing a hydrogen bond donor or acceptor, which is conducive to the recognition and binding of specific base sequences through the hydrogen bond; Third, it is best to have a positive or no negative charge on the side chain, which is conducive to the binding with the phosphoric acid backbone. The amino acids that meet these conditions are LYS, ARG, GLN, ASN and TYR.

2.12: Screening for a drug that binds to and stabilizes the helical structure of dìng, preventing conformational changes after they are encountered.

3.3: No, you cannot. The collagen triple helix is formed by folding the collagen in the precursor (pre-collagen) state, and after folding, it loses a section of the amino acid sequence, and the primary structure changes, so if the folded collagen is denatured, it is difficult to revert.

4.5：

253.2: These three effects are beneficial for HB to release bound oxygen to peripheral tissues when the partial pressure of oxygen and pH are low, which is essential for the function of HB to transport oxygen, but they are all based on the fact that HB has a quaternary structure, and since MB is composed of only one peptide chain and has no quaternary structure, it is impossible to have these three effects; In addition, MB's function is to store oxygen for muscle cells, so it does not need to have these three effects.

253.3: In humans, 2,3-bpg is stable dìng deoxyhemoglobin, while crocodiles lack this mechanism, but its hemoglobin likes to bind HCO3ˉ, and the idea of conducting deoxygenation is induced, so that high concentrations of CO2 are as stable as BPG in humans, and the binding of HCO3ˉ leaves H to help rebind hemoglobin and reduce pH. Therefore, with the production of CO2, oxygen is released.

253.6: (1) Phe, TRP, Val or TYR (2) Hydrophobic (3) HSP binds to them, preventing the binding between protein molecules to form precipitates between hydrophobic side chains and injure cells.

253.8: Determination of protein size and PI by SDS-PAGE and isoelectric aggregation, respectively; Northern blotting and RN blotting were used to detect whether it was expressed in hepatocytes; Its tertiary structure is determined using X-ray diffraction or NMR.

Chapter 5:

6.9: According to the codon and anticodon pairing, adhere to the principle of anti-parallelism and write the base sequence in the direction from 5 to 3, in addition, the ψ is pseudouridine, paired with A, it is not difficult to conclude that the codon sequence recognized by Gψc should be GAC.

7.3: Microorganisms living in the South Pole will contain more AT base pairs in the primary structure of the genome, and the tertiary structure is negative supercoiling, and the microorganisms living in volcanic craters should contain more GC base pairs in the primary structure of the genome, and the tertiary structure is likely to be positive supercoiled, so as to maintain the stability of their genomic DNA in a high temperature environment.

7.4: atatatatat

7.8: The denatured DNA buoyancy density increases, so the position will move downward.

253.15: The genomic DNA of microorganisms living in the Antarctic is rich in AT base pairs and forms a negative supercoil, as this facilitates the unstranding of DNA at low temperatures; The genomic DNA of microorganisms living in hot springs should be rich in GC base pairs and may form positive supercoiling, as this contributes to the stabilization of genomic DNA in high temperature environments.

253.16: Negative supercoiled DNA and positive supercoiled DNA are caused by insufficient and excessive entanglement of the two strands of DNA, respectively, and nuclease S1 specializes in hydrolyzing single-stranded nucleic acids, so when the covalent closed-loop negative supercoiled DNA and nuclease S1 are kept together, because the negative supercoiled covalent closed-loop DNA is in the solution and the relaxed DNA containing partial melting, nuclease S1 will hydrolyze the single-stranded part of the thawed loose DNA, resulting in a gap. This further tilts the equilibrium towards the flaccid type, and after a period of time, all negative supercoiled DNA is transformed into linear double-stranded DNA. However, positive supercoiled DNA does not form a flaccid DNA with a single-stranded region, so its structure remains unchanged.

Chapter 8: quiz2

Chapter 9: Q3, 4, 5, 8

Chapter 10: Q4, 6, 7

Chapter 11: Q1, 3, 4

Chapter 12: Q4, 5, 7

Chapter 13: Q1, 3, 5

Thought Questions (P253-254): Questions 9, 11, 12, 13.

Chapter 8:

2: The PKA of the HIS side chain is close to physiological pH, making it easy for it to participate in enzymatization as a proton donor or acceptor.

Chapter 9:

3: Hanes-wolff plotting, it uses [s]/v to [s] to plot, which avoids the influence of v error on the horizontal axis, so compared with the double reciprocal method and eadie-plotting method, the error of s-wolff plotting is small.

4: Irreversible inhibition decreases the effective concentration of the enzyme and leads to the decrease of zhìvmax, but does not affect the affinity between the enzyme and the substrate, and does not affect KM, so its dynamic characteristics are similar to non-competitive inhibition.

Group-specific inhibitors can covalently modify specific amino acid residues in the active center, and only care about whether there are amino acid residues that can be modified in the active center, regardless of what the three-dimensional structure of the active center is, so the specificity of their effect is the lowest.

5: Because these two inhibitors have the highest requirements for the specificity of the central structure of the enzyme's activity, that is, the most selective action, the use of them as primers causes the lowest side effects.

8: The larger the H, the stronger the positive synergy, the effect of the allosteric activator makes more enzymes in the reaction system become the R state, and the activation of the substrate is no longer needed, so the storage of the allosteric activator can lead to the decrease of the positive synergy of the allosteric enzyme, while the allosteric inhibitor is just the opposite, so the allosteric activator reduces H, and the allosteric inhibitor improves the □□.

Chapter 10:

4: Determine the properties of amino acid residues close to the binding site of the peptide substrate to see whether there is a catalytic triplet; The primary structure was determined, and the homology was investigated by comparing it with the known serine protease. Perform a series of kinetic pH changes to determine the PKA of key amino acid residues; use of serine protease-specific inhibitors and transition-state analogue inhibitors; Affinity labeling, chemical modifications, and site-directed mutagenesis can also be used.

6: Generalized acid-base catalysis, covalent catalysis of the unique anionic cavity to the carbon tetrahedral transition state. The anionic cavity contributed the most to the stability of the carbon tetrahedral transition state.

7: The PKA of a free cys sulfhydryl group is 8.4, but the same sulfhydryl gene in a protein molecule varies greatly depending on the microenvironment in which it is located, ranging from 5 to 10. The catalytic efficiency of thiol proteases is obviously related to the strength of the nucleophilic ability of thiol groups. The nucleophilic ability of sulfhydryl groups is related to whether it is ionized (loss of protons, dissociation), and the lower the PKA, the more likely it is to dissociate. Therefore, the optimal pH of different sulfhydryl proteases depends on the PKA of the sulfhydryl group in their active center.

As for the cys and Ser on some thiol proteases that are still active after substitution, it may be that its active center originally contains His and ASP, which can form a catalytic triplet with the introduced Ser.

Chapter 11:

1: The natural ribozymes that are still present in all living organisms are ribonuclease p and ribosomes; The ribozyme that is still present in all eukaryotes is U6-snRNA.

3: RNA in RNase P can be obtained by artificial synthesis or in vitro transcription, which can avoid protein contamination and see if it has catalytic activity.

4: According to the hypothesis of the RNA world, the first thing to do is to find out if there is RNA on this planet.

Chapter 12:

4: Theoretically, hydrophilic amino acid residues can be phosphorylated.

5: Trypsin, because other proteases are activated by the action of trypsin.

7: Hydrolytic activation, where natural transition-state analogues prevent premature activation of trypsin and inhibition of it by serpin as an inhibitory protein within trypsin-secreting cells.

Chapter 13:

1: Ethanol dehydrogenase can directly determine the activity of guònadh through the change of light absorption; Reverse transcriptase can be measured by measuring the radioactivity changes caused by the introduction of radiolabeled DNTP into DNA; Protein kinases are measured by the radioactivity of guò[γ-32p]-ATP to phosphorylated proteins.

3: Compare the results of electrophoresis on native gels and SDS-gels to see that there is no change in bands.

5: Theoretically, any hydrophilic amino acid is OK because their side chain groups are chemically reactive.

Thought Questions (P253-254): Questions 9, 11, 12, 13.

9: It should be a trypsin inhibitor, because the other two proteases are converted from zymogen to enzyme by trypsin activation

11: Coenzymes bind to enzymes so strongly that they are difficult to isolate by gentle methods such as dialysis and ultrafiltration, which is not the case with coenzymes. The coenzymes or cogroups that contain adenosine are: Coenzyme I, Coenzyme II, FAD and CoA. This can be used as evidence for the RNA world hypothesis.