"You have run both non-denaturing and denaturing samples of your purified fluorescent protein on the SDS-PAGE...
"You have run both non-denaturing and denaturing samples of your purified fluorescent protein on the SDS-PAGE gel. When do you expect to see a difference between the two samples? Briefly explain."
Hey!! if anyone could help me with this question I would greatly appreciate it. It is based on my cell bio lab and I don't understand the question. Thankyou in advance!!
Homework Answers
In SDS PAGE, SDS denatures proteins by wrapping. around the polypeptide backbone. By heating the sample in the range of 70-100°C in the presence of excess SDS and thiol reagent, disulfide bonds are broken or cleaved, and the protein is fully dissociated into its subunits, so in both cases protein is denatured in SDS PAGE but if you are running known protein sample you can analyze it with reference.
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"You have run both non-denaturing and denaturing samples of your
purified fluorescent protein on the SDS-PAGE...
You have purified a His-tagged protein expressed in human cell culture by Ni2+ affinity chromatography. You...
You have purified a His-tagged protein expressed in human cell
culture by Ni2+ affinity
chromatography.
You have purified a His-tagged protein expressed in human cell culture by Ni^2+ affinity chromatography. When you run the sample on an SDS PAGE gel in the absence of reductant you observe a single band; in the presence of reductant, however, you observe two bands. If you treat with reductant and pour the products over the nickel column, you detect one band in the flow-through...
Question 6 You and your G-protein coupled receptors o You have encountered G-protein coupled receptors in...
Question 6 You and your G-protein coupled receptors o You have encountered G-protein coupled receptors in Intro Bio I lecture (or soon will), and will encounter them briefly again in Intro Bio II. You will spend 2 or 3 lectures on them in Cell Bio. They are integral (transmembrane) proteins embedded in the plasma membrane of every eukaryotic cell in your body. We will treat them as being 50 Angstroms in diameter. Considering the plasma membrane scaled up to the...
You find yourself setting up another Western blot, however. today you are preparing the samples t...
You find yourself setting up another Western blot, however. today you are preparing the samples to load onto the SDS-PAGE gel. You are using a 4-20%SDS-PAGE gel that can hold a maximum of 30 μしper 1 You remember from you so decided to prepare each sample with a final volume of 25 uL. However, no one is perfect at pipetting, so make sure each sample is made at 1.5X the total volume needed per well. The protein you are interested...
can anyone help me with this cell bio question? In Figure 2 part A (Non-denaturing). Lane 1 NEP( E403C) , 2 = WT...
can anyone help me with this cell bio
question?
In Figure 2 part A (Non-denaturing). Lane 1 NEP( E403C) , 2 = WT ECE, , 4= WT= NEP, 9= ECE(C416E). Figure 2 part B. The same as part A, but under denaturing conditions. (note; for simplicity, many lanes are ignored) 1 2 3 5 67 89 10 a -213 -123 -85 -50 b kDa -213 -123 85 -50 Figure 2 Immunoblot analyses of wild-type and cysteine mulants of NEP and...
Why are proteins heat denatured prior to analysis in SDS-PAGE? Select all answers that apply. Denaturation...
Why are proteins heat denatured prior to analysis in SDS-PAGE? Select all answers that apply. Denaturation of the protein is necessary so that proteins run proportionally to their size, based on the interaction of SDS with the unfolded protein. Heat is used to hydrolyze the peptide bonds of the protein. The heat step allows the proteins to unfold, enabling the protein chain to be coated with SDS molecules. Heat is used to hydrolyze disulfide bonds. QUESTION 2 1.5 points Saved...
In your previous prac session you digested your POTC-A plasmid DNA with 3 enzyme mixes (AB...
In your previous prac session you digested your POTC-A plasmid DNA with 3 enzyme mixes (AB and C). One mix contained the restriction endonuclease Kpnl alone, another contained both del and Not and another contained Sphi and Nhe, but you don't know which was which yet. The sites where these enzymes cut POTC-A are indicated on the plasmid map at the top of the page. Calculate the sizes of the DNA fragments that you would expect to see on your...
You have purified a His-tagged protein expressed in human cell
culture by Ni2+ affinity
chromatography.
You have purified a His-tagged protein expressed in human cell culture by Ni^2+ affinity chromatography. When you run the sample on an SDS PAGE gel in the absence of reductant you observe a single band; in the presence of reductant, however, you observe two bands. If you treat with reductant and pour the products over the nickel column, you detect one band in the flow-through...
Question 6 You and your G-protein coupled receptors o You have encountered G-protein coupled receptors in Intro Bio I lecture (or soon will), and will encounter them briefly again in Intro Bio II. You will spend 2 or 3 lectures on them in Cell Bio. They are integral (transmembrane) proteins embedded in the plasma membrane of every eukaryotic cell in your body. We will treat them as being 50 Angstroms in diameter. Considering the plasma membrane scaled up to the...
You find yourself setting up another Western blot, however. today you are preparing the samples to load onto the SDS-PAGE gel. You are using a 4-20%SDS-PAGE gel that can hold a maximum of 30 μしper 1 You remember from you so decided to prepare each sample with a final volume of 25 uL. However, no one is perfect at pipetting, so make sure each sample is made at 1.5X the total volume needed per well. The protein you are interested...
can anyone help me with this cell bio
question?
In Figure 2 part A (Non-denaturing). Lane 1 NEP( E403C) , 2 = WT ECE, , 4= WT= NEP, 9= ECE(C416E). Figure 2 part B. The same as part A, but under denaturing conditions. (note; for simplicity, many lanes are ignored) 1 2 3 5 67 89 10 a -213 -123 -85 -50 b kDa -213 -123 85 -50 Figure 2 Immunoblot analyses of wild-type and cysteine mulants of NEP and...
Why are proteins heat denatured prior to analysis in SDS-PAGE? Select all answers that apply. Denaturation of the protein is necessary so that proteins run proportionally to their size, based on the interaction of SDS with the unfolded protein. Heat is used to hydrolyze the peptide bonds of the protein. The heat step allows the proteins to unfold, enabling the protein chain to be coated with SDS molecules. Heat is used to hydrolyze disulfide bonds. QUESTION 2 1.5 points Saved...
In your previous prac session you digested your POTC-A plasmid DNA with 3 enzyme mixes (AB and C). One mix contained the restriction endonuclease Kpnl alone, another contained both del and Not and another contained Sphi and Nhe, but you don't know which was which yet. The sites where these enzymes cut POTC-A are indicated on the plasmid map at the top of the page. Calculate the sizes of the DNA fragments that you would expect to see on your...
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