Proteins thread?
Any biochemists, biophysicists etc etc working with proteins?
I work with crystallography and NMR. I recently made a segmentally 15N-labelled protein on its IDR using sortase from S. Aureus, ready to do some HSQC spectra on it.
math fag here. how do you retards know if the shit is folded correctly
>>8069233
NMR spectroscopy is a method that exploits the resonance of the nuclei and can provide atomic resolution information to help determine structure and dynamics of a protein. By applying a powerful magnetic field on the protein you can get data which reveals the fold of the protein
>>8069233
leave the real science to the big boys
>>8069237
are the values discrete and widely separated?
How do you tell if its exact or you just made a version of it that looked similar
>>8069306
This is the dataset from a 2D HSQC (hetero-nuclear single quantum correlation) showing a peak for each of the amino acid (AA) residues in the protein. Each AA will have a unique environment depending on its chemical envronment, which affects the chemical shift (the frequency independt of the magnetic field applied). When added a ligand or something else, the spectra will change if the ligand binds, as the environment changes around some of the residues.
This is a simple NMR experiment, as they often are conducted as 3D experiments, adding another bond correlation to the spectra.
>>8069306
Not OP, structural biologist here.
The values are mostly separated widely enough, but it doesn't matter when they're not because they're proportional. Meaning if you have two atoms that cause the same data point, it'll have twice the amplitude.
The cool thing about NMR is that it's incredibly sensitive, which also means you can see the dynamic aspect as well. i.e. you can see that a fold isn't rigid, it has more or less freedom. You just look for the general conserved structure when there is one.
Some proteins don't even have a well-defined structure, we just call them intrinsically disordered proteins.
>>8069306
If you mean, how i can be sure that the structure is the native structure as in vivo (in the cell)?
It is actually possible to conduct an in-cell NMR experiment. Only spin-1/2 nuclei show on an NMR spectra - they have a magnetic spin moment. These spin-1/2 nuclei are 15N or 13C etc. It is possible to specificly label a protein in E. Coli, making the cell overexpress your protein in very high yields, making it possible to perform NMR experiments on living cells! Very cool - and shuts up in-vitro hating biologists.
>>8069363
Yes but how can you be sure that the extremely magnetic environment in which you've placed your protein doesn't affect its conformation?
Please do a control NMR experiment without the magnetic fields.
- Reviewer 2
>>8069352
>skærmbillede
Er du biokemiker ved KU?
>>8069357
this is the exact reason I didnt do this stuff. I like my things well defined and exact. Stuff like this gives me brain aneurysm
>>8069385
Just think of it as a set of all possible conformations.
You have a protein, the positions of its atoms is a function of its own composition and everything around it. Defining it as a mathematical function is fucking retarded but that's what it is, really.
>>8069373
Ja, det er jeg
>>8069405
Haha, 4chan er det sidste sted jeg ville forvente at finde andre biokemikere.
Er selv dog bare en lowly 2. års noob.
Er det proteiner man skal specialisere sig i hvis man vil have 300k starting?
