Secondary Structure Inspection: Use the tools you learned for secondary structur
ID: 252740 • Letter: S
Question
Secondary Structure Inspection: Use the tools you learned for secondary structure determination to examine what secondary structures are present in this protein. Discuss your selections with appropriate reasoning (and pictures, if useful) in your lab notebook. ExPASy Bioinformatics Tools: You will be using ExPASy to calculate molecular weight, isoelectric point and extinction coefficient using the protparam feature (http://ca.expasy.org/tools/protparam.html). The tools on the ExPASy website require a sequence, which can be typed or pasted into the search boxes. Before you do this, delete all chains in the 2GSN PDB file except the first chain. To do this, right click on the other chains and select "Delete This Chain" in the pop-up menu. Then, copy the sequence in fasta format and paste it into the search box in the ExPASy protparam page. You can copy the sequence from the Sequence Viewer window of MOE in several formats using the Edit->Copy SEQ command. The first line of the sequence (starting with >) is a comment line that should be deleted before clicking 'Compute Parameters' in the ExPASy window. Discuss the relationship between the isoelectric point and the relative numbers of residues with acidic and basic sidechains in your lab report Data Analysis What type of ion exchange chromatography would you propose to use if trying to purify your protein from many other proteins? Justify your choice based on its pl Use the calculated molecular weight of the protein to determine the amount of your proteirn required to make 5 mL of a 10 HM solution for further fictitious spectroscopic analysis and show your math in your lab notebook. What absorbance would you predict for this solution at 280 nm based on the calculated extinction coefficient (using the Beer-Lambert law)? .Why is the extinction coefficient determined at 280 nm? What chemical entities contribute to a maximum absorbance at this absorbance?Explanation / Answer
1- Given that the pI of the protein is 4.6. It means that the protein has overall negative charge. In order to purify the protein from the pool of proteins, we need to use cation resin, which can bind to the negatively charged protein.
2- the Molecular weight of the protein is 2803kDa.
Molarity is given by the number of moles in per litre of solution.
10micromol = Weight / Mw *1000/ volume in ml
10 *10-6 = Weight / 2803272 * 1000/ 5
= 140 mg.
3- Given that the extinction coefficient of the protein is 418560 as given. For measuring extinction coefficient we use the cystines residues.
Absorbance = ? c L
L = path length = 1 cm
c = 10micromol.
418560 * 10*10-6 *1
0.418560.
4- amino acids such as tyrosine, tryptophan and cystine absorb at 280nm. So we measure extinction coefficient at 280 nm.
5. At 280nm maximum absorbance is shown by tryptophan, which is 5500 Followed by tyrosine which is 1490 and then by cystine which is 120.
Related Questions
drjack9650@gmail.com
Navigate
Integrity-first tutoring: explanations and feedback only — we do not complete graded work. Learn more.