IPTG. Explain the mechanistic steps whether the expression of beta-galactosidase (Z) and permease (Y) are inducible (I) or noninducible (N) or constitutive (C).
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- For the genotypes and conditions (lactose present or absent) shown in the following Table 2, predict whether the expression of beta-galactosidase is inducible (I), noninducible (N) or constitutive (C). Explain your reason. Table 2 Genotype I*p*O°z* Condition (i) No lactose (ii) I*P*O*Z+ Lactose I*P-0*Z* (iii) (iv) Lactose ISP+O°Z+ Lactose (v) Ip*o°Z+ No lactoseFor the lac genotypes of Escherichia coli shown in the following Table 1, predict the expression of beta-galactosidase (Z) and permease (Y) is inducible or noninducible or constitutive. Explain your answer. Table 1 Genotype I-P+O+Z+Y+ (i) (ii) I+P+OCZ+Y+ (iii) ISP+O+Z+Y+ (iv) I+P+O+Z+Y-//I+P-O+Z+Y+ (v) ISP+OcZ+Y+//I-P+O+Z+Y- Condition No lactose No lactose lactose lactose No lactoseThe lac genotypes are as shown below: P+OcZ-Y+A+// P¯O+Z+Y+A+ (i) The lac operon consists of three structural genes, lacZ, lacY and lacA. Which structural genes are involved in lactose metabolism? Explain. (ii) Draw and explain how lactose repress the gene expression in lac IS/I- heterozygote. (iii) What is the function of the promoter in the bacterial operon?
- The first 32 amino acids from the N terminus of the protein bovine angiogenin were determined by Edman degradation and have the sequence:AQDDYRYIHFLTQHYDAKPKGRNDEYCFNMMK(a) Identify the sites of cleavage during trypsin-catalyzed hydrolysis of this protein.(b) What are the cleavage sites using chymotrypsin?For each of the E. coli strains that follow, indicate theeffect of the genotype on the expression of the trpEand trpC genes in the presence or absence of tryptophan. [In the wild type (R+ P+ o+ att+ trpE+ trpC+),trpC and trpE are fully repressed in the presence oftryptophan and are fully expressed in the absence oftryptophan.]R = repressor gene; Rnproduct cannot bind tryptophan; R− product cannot bind operatoro = operator for the trp operon; o− cannot bind repressoratt = attenuator; att− is a deletion of the attenuatorP = promoter; P− is a deletion of the trp operonpromotertrpE− and trpC− are null (loss-of-function) mutationsa. R+ P− o+ att+ trpE+ trpC+b. R− P+ o+ att+ trpE+ trpC+c. RnP+ o+ att+ trpE+ trpC+d. R− P+ o+ att− trpE+ trpC+e. R+ P+ o− att+ trpE+ trpC−/R− P+ o+ att+trpE− trpC+f. R+ P− o+ att+ trpE+ trpC−/R− P+ o+ att+trpE− trpC+g. R+ P+ o− att− trpE+ trpC−/R− P+ o− att+trpE− trpC+In the "biochemical assay of b-galactosidase activity", what is o-nitrophenyl-b-D-galactosidase (ONPG) used for? O When ONPG is cleaved by b-galactosidase, we are able to assay b-galactosidase activity. O It provides essential nutrients for bacterial growth. It partially disrupts the cell membrane to allow cellular proteins to diffuse out of the cell. O ONPG cleaves the b-galactosidase that is made by the lac operon so we can see how much activity there is in the cell. ONPG cleaves galactose so we can measure how much lactose there is in the cell.
- Explain how the following mutations would affect transcription of the yeast GAL1 gene in the presence of galactose. (a) A deletion within the GAL4 gene that removes the region encoding amino acids 1 to 100. (b) A deletion of the entire GAL3 gene. (c) A mutation within the GAL80 gene that blocks the ability of Gal80 protein to interact with Gal3p. (d) A deletion of one of the four UASG elements upstream from the GAL1 gene. (e) A point mutation in the GAL1 core promoter that alters the sequence of the TATA box.Clary Fray used the pET vector system to express her prokaryotic amylase enzyme. She added peptone into her culture broth of BL21(DE3) Escherichia coli strain to induce protein expression. At the end of the experiment, she discovered that her protein was not expressed. She repeated three more times but her protein of interest was still not produced. (i) (ii) (iii) (iv) (v) Explain the reason why Clary failed to obtain her protein of interest and suggest a solution to troubleshoot this problem. Clary plans to express her protein along with a polyhistidine-tag. Explain the importance of His-tag in protein work. Is DH5a Escherichia coli suitable to propagate the plasmid before protein expression? Besides heat shock method, elaborate another procedure Clary could utilize to transform the recombinant pET vector into the host cell. If her supervisor instructs her to express a gene from gold fish (Carassius auratus), is the expression system she is using now suitable for this experiment?…I have this strain of e coli. Is P+ o+ Z+ Y+ / I- P+ oC Z- Y+ Will beta-galactosidase and permease be expressed? If they are will they be inducible or constitutive?
- CTP synthetase catalyzes the glutamine-dependent conversion of UTP to CTP. The enzyme is allosterically inhibited by the product, CTP. Mammalian cells defective in this allosteric inhibition are found to have a complex phenotype: They require thymidine in the growth medium, they have unbalanced nucleotide pools, and they have an elevated spontaneous mutation rate. Explain the likely basis for these observations.Consider the following simple regulatory pathways. Assume the full pathway is shown. A- E- B- F- C- G- D- 1 A H- 2 B || L You identify several null mutations (a complete deletion of the gene). For each mutant (indicated with a - sign), determine whether the final product (I, J, K or L) is inducible, uninducible, or constitutive. 3 C 4 D- [Choose ] [Choose ] [Choose ] [Choose ] [Choose ] [Choose ] [Choose ] E [Choose ] F G I H || J KFor each of the following genotypes, explain how mutation (identified by a (-) will affect the organism grown in lactose medium. Indicate whether a) B-galactosidase will be synthesized or not, b) synthesis of B-galactosidase is inducible (1) or constitutive (C) and c) growth of the organism will occur or not. i. I+P+0-Z+Y+A+/l+P+O+Z-Y-A- ii. I-P+0+Z+Y+A+/I+P-O+Z+Y+A+