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Questions related from Prem Prakash
Two different proteins which are well known for forming a hexamer ring by making electrostatic interaction among its heterodimers, reported in yeast and the structure is also available in PDB. I...
18 July 2022 3,668 5 View
Dear all, I have started performing an experiment to determine the Kd value of an exonuclease enzyme using 5'-FAM labelled Oligo substrates using Fluorescence polarization. As this enzyme uses...
23 July 2021 1,587 16 View
It is very much confusing when some papers use this term "apparent Km of an enzyme even if they use single substrate ? Not at all clear ! Please help me understanding this. Thanks in advance.
28 September 2020 3,729 7 View
I am planning to make the wild type enzyme pH tolerant (acidic) without affecting the activity of the native enzyme. Here i am thinking to introduce some residues like lysine or ariginine in place...
27 September 2017 1,453 4 View
I have done the flourescence spectrometry experiment with my protein and ligand. Keeping the protein concentration constant with increasing concentration of the ligand (here NADH). I saw there is...
04 June 2015 8,238 4 View
I have done site directed mutagensis by designing the primers at the mutation site, and whole plasmid amplification was performed using Hifidelity enzyme XT-20 (which can amplify up to 20 kb). The...
17 December 2014 945 12 View
As I have purified my protein and set crystallization screen and I tried various way to optimize the crystal quality, such as seeding, changing pH, PEG concentration, salt concentration and...
20 May 2014 4,426 9 View
Why it is problematic during molecular replacement or any other simulation, is there any comment ?
01 December 2013 8,688 8 View
Does anyone have a very clear cut explanation for the above terms? I am unable to understand from the sources in Google. And why a detector is so important in any data reduction program ?
09 October 2013 9,037 3 View
If a (purified) protein of an unknown gene interacts with an antibody, is it possible to get the nucleotide sequence without protein sequencing? Does any body have any explanation for this ?
09 June 2013 7,401 10 View
I have to do an enzyme kinetic assay using NADPH as a substrate (10 mM) and in one shot I cannot finish my kinetic assay with all the fractions collected at each step of the purification. By that...
05 May 2013 3,203 6 View
I have purified my protein and want to estimate the amount. The plot I am getting is half of the value my senior is getting. The difference is I am using a kit (Genei TM) for Bradford's assay. But...
23 January 2013 7,574 24 View
Can I store cell lysate in -20 degree ( the protein for structural determination) freezer for one or two days?
02 January 2013 5,125 1 View
And if mutation has been done (in protein gene) and then there is no activity found with enzyme how one can monitor the function after mutation. Please explain both question if its possible.
14 May 2012 5,616 2 View
Please suggest. What should be the pH of this when prepared?
18 March 2012 7,095 5 View
Can anybody explain how to proceed with cell culture of this cell line? Thank you!
14 March 2012 464 15 View
As I have to suggest my profs.about this. Please give your opinions !! The subjects should be apart from Bioinformatics, Biophysics, and genetic-eng. enzymology and immunology.
07 March 2012 2,265 6 View
Dear all, Apologies for an off topic. I am curious to know if someone wants to pursue a second PhD in order to switch into a closely related field from his/her first PhD, would that be considered...
01 January 1970 8,976 4 View
Dear all, Greetings ! I am a total newbie in enzymology as I started working on this very field couple of months ago. I am working on a nuclease which degrades oligomers of nucleotide from 3'...
01 January 1970 200 17 View
I have infected my insect cells (SF-21), with P4 viral titre. Now the protocol suggest that it should be harvested before lysis. Can anyone suggest me how I will figure out when to harvest after...
01 January 1970 5,722 6 View
Dear all, I am trying to check the interaction between two of my proteins of interest. However before trying any biophysical technique such as SPR/ITC/MST etc, I used normal superdex-200 gel...
01 January 1970 4,928 3 View
As I read that, designing assay for cysteine protease is a bit challenging compare to other proteases, I am wondering why it is so ? Many research articles follows fluorescence based assay for...
01 January 1970 8,433 4 View
Dear All, I am trying to express a large protein (220 kDa) with a GST tag. I performed Western Blot to check for the protein of interest. Now after using monoclonal antibody (Mouse Ab), I have...
01 January 1970 5,012 9 View
Dear All, I am trying to express and purify a protein of MW 220 kDa (full length). It has an extra GST tag in it. The strain I am using is Rossetta (so issue of rare codon is ruled out). Now,...
01 January 1970 1,612 5 View
Dear All, My initial or original predicted model of protein has all the amino acid sequences in the polypeptide chain. But when I docked this structure with its partner protein, the resultant...
01 January 1970 2,165 9 View
Dear all, I got results after performing an enzyme kinetics with 4 different substrate the kcat of B and D is respectively two times lower than A and C. However, I am seeing the higher Km of A...
01 January 1970 8,749 4 View
Dear all, I am looking for a commercially available pull down assay kit that has conducive buffer conditions and high affinity for the antibody conjugation. My protein of interest (bait) shows a...
01 January 1970 3,496 5 View
