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1. You are working on identifying novel proteins associated with the plasma membrane and you isolate XYZ1. Protein sequencing shows XYZ1 is comprised of: MGASSCSELRSSAFALSERSDKSSAVLLIVLGVMAVGIYLLVTMVLIIGTSANRASGSKD SESMSTLECAAPNDSAA a. Based on the sequence. Do you predict this is a peripheral membrane protein or an integral membrane protein? (Hint: use the Hydropathy plot generator on Expasy using default settings to help answer this question.) b. You decide to conduct an experiment to further assess how the protein is connected to the PM. The experiment is conducted by either 1) treating intact cells with chymotrypsin, 2) by disrupting the plasma membrane such that chymotrypsin has access to both sides of the plasma membrane, or 3) by using detergents to remove all lipid bilayers and treating the proteins such that chymotrypsin will have access to -Chym. Cond. 1 +Chym. Cond. 2 Cond. 3 the entire length of the protein. The treatments are run on an SDS-PAGE gel (together with an untreated control) and immunoblotted using anti-XYZ1 antibodies. i. What amino acid residues does chymotrypsin proteolytically cleave after? ii. Is the protein peripheral/integral? How do you know? What would the results have looked like if it were the opposite result? iii. Digestion at Y-39 (right in the middle of the XYZ1) does not happen until condition 3. Based on your answer in (ii) what might be preventing digestion at this site? iv. You conduct a proteomics study and find that XYZ1 is palmitoylated on Cys-69. How might you further explore the importance of this modification on XYZ1 function?

1. You are working on identifying novel proteins associated with the plasma membrane and you isolate XYZ1. Protein sequencing shows XYZ1 is comprised of: MGASSCSELRSSAFALSERSDKSSAVLLIVLGVMAVGIYLLVTMVLIIGTSANRASGSKD SESMSTLECAAPNDSAA a. Based on the sequence. Do you predict this is a peripheral membrane protein or an integral membrane protein? (Hint: use the Hydropathy plot generator on Expasy using default settings to help answer this question.) b. You decide to conduct an experiment to further assess how the protein is connected to the PM. The experiment is conducted by either 1) treating intact cells with chymotrypsin, 2) by disrupting the plasma membrane such that chymotrypsin has access to both sides of the plasma membrane, or 3) by using detergents to remove all lipid bilayers and treating the proteins such that chymotrypsin will have access to -Chym. Cond. 1 +Chym. Cond. 2 Cond. 3 the entire length of the protein. The treatments are run on an SDS-PAGE gel (together with an untreated control) and immunoblotted using anti-XYZ1 antibodies. i. What amino acid residues does chymotrypsin proteolytically cleave after? ii. Is the protein peripheral/integral? How do you know? What would the results have looked like if it were the opposite result? iii. Digestion at Y-39 (right in the middle of the XYZ1) does not happen until condition 3. Based on your answer in (ii) what might be preventing digestion at this site? iv. You conduct a proteomics study and find that XYZ1 is palmitoylated on Cys-69. How might you further explore the importance of this modification on XYZ1 function?

Biology 2e
Biology 2e
2nd Edition
ISBN:9781947172517
Author:Matthew Douglas, Jung Choi, Mary Ann Clark
Publisher:Matthew Douglas, Jung Choi, Mary Ann Clark
Chapter5: Structure And Function Of Plasma Membranes
Section: Chapter Questions
Problem 7RQ: A scientist compares the plasma membrane composition of an animal from the Mediterranean coast with...
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1. You are working on identifying novel proteins associated with the plasma membrane and you isolate XYZ1. Protein sequencing shows XYZ1 is comprised of: MGASSCSELRSSAFALSERSDKSSAVLLIVLGVMAVGIYLLVTMVLIIGTSANRASGSKD SESMSTLECAAPNDSAA a. Based on the sequence. Do you predict this is a peripheral membrane protein or an integral membrane protein? (Hint: use the Hydropathy plot generator on Expasy using default settings to help answer this question.) b. You decide to conduct an experiment to further assess how the protein is connected to the PM. The experiment is conducted by either 1) treating intact cells with chymotrypsin, 2) by disrupting the plasma membrane such that chymotrypsin has access to both sides of the plasma membrane, or 3) by using detergents to remove all lipid bilayers and treating the proteins such that chymotrypsin will have access to -Chym. Cond. 1 - +Chym. Cond. 2 Cond. 3 the entire length of the protein. The treatments are run on an SDS-PAGE gel (together with an untreated control) and immunoblotted using anti-XYZ1 antibodies. i. What amino acid residues does chymotrypsin proteolytically cleave after? ii. Is the protein peripheral/integral? How do you know? What would the results have looked like if it were the opposite result? iii. Digestion at Y-39 (right in the middle of the XYZ1) does not happen until condition 3. Based on your answer in (ii) what might be preventing digestion at this site? iv. You conduct a proteomics study and find that XYZ1 is palmitoylated on Cys-69. How might you further explore the importance of this modification on XYZ1 function?
Transcribed Image Text:1. You are working on identifying novel proteins associated with the plasma membrane and you isolate XYZ1. Protein sequencing shows XYZ1 is comprised of: MGASSCSELRSSAFALSERSDKSSAVLLIVLGVMAVGIYLLVTMVLIIGTSANRASGSKD SESMSTLECAAPNDSAA a. Based on the sequence. Do you predict this is a peripheral membrane protein or an integral membrane protein? (Hint: use the Hydropathy plot generator on Expasy using default settings to help answer this question.) b. You decide to conduct an experiment to further assess how the protein is connected to the PM. The experiment is conducted by either 1) treating intact cells with chymotrypsin, 2) by disrupting the plasma membrane such that chymotrypsin has access to both sides of the plasma membrane, or 3) by using detergents to remove all lipid bilayers and treating the proteins such that chymotrypsin will have access to -Chym. Cond. 1 - +Chym. Cond. 2 Cond. 3 the entire length of the protein. The treatments are run on an SDS-PAGE gel (together with an untreated control) and immunoblotted using anti-XYZ1 antibodies. i. What amino acid residues does chymotrypsin proteolytically cleave after? ii. Is the protein peripheral/integral? How do you know? What would the results have looked like if it were the opposite result? iii. Digestion at Y-39 (right in the middle of the XYZ1) does not happen until condition 3. Based on your answer in (ii) what might be preventing digestion at this site? iv. You conduct a proteomics study and find that XYZ1 is palmitoylated on Cys-69. How might you further explore the importance of this modification on XYZ1 function?
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