Hi everyone!
While doing research for my master's thesis I have encountered something that I have not been able to find the answer to. I am using Western blotting to compare TGF-beta expression between lysate and conditioned media, and the antibody I am using identifies three forms of the protein: monomer, dimer, and latent.
The TGF-beta homodimer is known to be biologically active, and I understand that the latent protein is inactive because a latency associated peptide (also a dimer) prevents TGF-beta from interacting with the receptor.
My question is this -- would the monomeric form be active? Or is the receptor sensitive such that only the dimer is involved in cytokine signaling? Furthermore, if the protein is formed as a dimer and exported to the extracellular matrix in this form as literature suggests, what catalyzes the formation of TGF-beta monomers?
The 3.4 Å structure of cytochrome c oxidase from the hyperthermophilic bacterium Aquifex aeolicus (AaCcO) has been solved. The molecular mechanism that AaCcO uses involves both cytochrome c and quinol as electron donors through the native quinol molecules (NQs) bound at the dimeric interface.
Abstract
The heme‐copper oxidase superfamily comprises cytochrome c and ubiquinol oxidases. These enzymes catalyze the transfer of electrons from different electron donors onto molecular oxygen. A B‐family cytochrome c oxidase from the hyperthermophilic bacterium Aquifex aeolicus was discovered previously to be able to use both cytochrome c and naphthoquinol as electron donors. Its molecular mechanism as well as the evolutionary significance are yet unknown. Here we solved its 3.4 Å resolution electron cryo‐microscopic structure and discovered a novel dimeric structure mediated by subunit I (CoxA2) that would be essential for naphthoquinol binding and oxidation. The unique structural features in both proton and oxygen pathways suggest an evolutionary adaptation of this oxidase to its hyperthermophilic environment. Our results add a new conceptual understanding of structural variation of cytochrome c oxidases in different species.
https://ift.tt/33lWgzR
Just making sure I have this down, so someone please feel free to correct me if I am wrong:
Native PAGE separates proteins according to mass and charge. Additionally, since there is no reducing agent, any disulfide-linked protein will not separate into its constituent subunits. Thus, regardless of whether or not a dimer contains disulfide linkages, a homodimer will appear as one line, and a heterodimer will appear as one line. Basically, both dimers will run as monomers.
Non-reducing SDS-PAGE denatures proteins (removes their quaternary structure) and assigns them all a uniform negative charge so that proteins can be separated solely on the basis of mass. The results would be the same as for native PAGE, regardless of whether or not the proteins contain disulfide linkages.
Reducing SDS-PAGE uses a reducing agent (β-mercaptoethanol) to reduce disulfide linkages into their constituent sulfhydryl groups, thus allowing for the separation of disulfide-linked dimer into their constituent subunits. Thus, a disulfide-linked homodimer would appear as one large band (because the subunits, which are identical, have the same mass). For example, a disulfide-linked 80 kDa homodimer would show a single large band at 40 kDa. In contrast, a disulfide-linked heterodimer would appear as two distinct bands since the subunits are different from each other and thus have different masses. For example, a disulfide-linked 80 kDa heterodimer would show one band at 50 kDa and one band at 30 kDa (or any two combinations that add up to 80, depending on the passage of course).
KRAS: Paramagnetic relaxation enhancement (PRE) NMR spectroscopy approaches combined with a nanodisc system reveal distinct structures of membrane‐associated homodimers of GTP‐ versus GDP‐bound KRAS4B. Both structures dimerize via a α4–α5 interface, but differ in the relative orientation of the protomers. KRAS4B‐GTP dimerization increases the accessibility of the effector binding site for RAF kinases.
Abstract
KRAS homo‐dimerization has been implicated in the activation of RAF kinases, however, the mechanism and structural basis remain elusive. We developed a system to study KRAS dimerization on nanodiscs using paramagnetic relaxation enhancement (PRE) NMR spectroscopy, and determined distinct structures of membrane‐anchored KRAS dimers in the active GTP‐ and inactive GDP‐loaded states. Both dimerize through an α4–α5 interface, but the relative orientation of the protomers and their contacts differ substantially. Dimerization of KRAS‐GTP, stabilized by electrostatic interactions between R135 and E168, favors an orientation on the membrane that promotes accessibility of the effector‐binding site. Remarkably, “cross”‐dimerization between GTP‐ and GDP‐bound KRAS molecules is unfavorable. These models provide a platform to elucidate the structural basis of RAF activation by RAS and to develop inhibitors that can disrupt the KRAS dimerization. The methodology is applicable to many other farnesylated small GTPases.
https://ift.tt/39vJYFT
Hey guys, I was studying NSAID pharmacology and this paragraph on G&G's got me stumped.
Here's the paragraph
>While the functional COX enzymes are sequence homodimers, they are configured as conformational heterodimers in which one of the monomers functions as the catalytic subunit with heme bound and the other, without heme, serves as the allosteric subunit.
What I got from it is this: COX enzymes are made up of two monomers. The aminoacid sequence in both monomers is the same (homodimers) but they are functionally (?) different; one catalyses the conversion of arachidonic acid to PGG2 while the other one serves as an allosteric subunit.
Did I get it right? If not I'd really appreciate your help.
Thanks!
Hi all, I can't seem to wrap my head around this UEarth question and the percentage breakdown for the different answers makes it seem questionable. The passage and questioned are outlined below (changed values and modified actual question to avoid copyright issues):
A homodimer protein dissociates into its monomers. Each monomer is 200-residues long. How many mRNA nucleotides must be translated to form the fully-functional protein?
I answered 600, since each residue requires 3 nucleotides in a codon and the fully functional protein is a homodimer (2 copies of a monomer).
They listed the correct answer and explanation as 1200 nucleotides.
I'm not sure why this is correct since the two monomers require the same 600 nucleotides. My only guess is that reading the question literally, the "how many nucleotides must be translated" doesn't specify unique nucleotides, so we answer with a literal number of nucleotides that were translated, and not individual nucleotides.
Sounds like 2 identical heterodimers are homodimers????
I am looking for a website that in a structured manner, with the assistance of infographics, can visually display how for example certain cells are interconnected with each other. 3D, 2D, charts, pie-charts etc.
Let us take the 5HT2-A receptor which has a complex range of interactions with the 5-HT1A, GABAA, adenosine A1, AMPA, mGluR2/3, mGlu5, and OX2 receptors. In the rat cerebellum, the protein has also been found in the Golgi cells of the granular layer, and in the Purkinje cells.
https://en.wikipedia.org/wiki/5-HT2A_receptor
A good site already exists that describes economic complexity - the observatory for economic complexity.
https://atlas.media.mit.edu/en/
It does not have to be super-complicated with a lot of interactions between proteins, but mainly how different receptors interact with each other.
I don't want to step on anybody's toes here, but the amount of non-dad jokes here in this subreddit really annoys me. First of all, dad jokes CAN be NSFW, it clearly says so in the sub rules. Secondly, it doesn't automatically make it a dad joke if it's from a conversation between you and your child. Most importantly, the jokes that your CHILDREN tell YOU are not dad jokes. The point of a dad joke is that it's so cheesy only a dad who's trying to be funny would make such a joke. That's it. They are stupid plays on words, lame puns and so on. There has to be a clever pun or wordplay for it to be considered a dad joke.
Again, to all the fellow dads, I apologise if I'm sounding too harsh. But I just needed to get it off my chest.
Do your worst!
I'm surprised it hasn't decade.
For context I'm a Refuse Driver (Garbage man) & today I was on food waste. After I'd tipped I was checking the wagon for any defects when I spotted a lone pea balanced on the lifts.
I said "hey look, an escaPEA"
No one near me but it didn't half make me laugh for a good hour or so!
Edit: I can't believe how much this has blown up. Thank you everyone I've had a blast reading through the replies 😂
It really does, I swear!
SDS page- WILL separate proteins by disrupting covalent bonds, but not disulfide. So if you have a homodimer composed of 25kda monomers, under SDS page the covalent bonds break and you just have the monomers traveling separately, each 25kda.
Under non reducing, disulfide bonds do not break. So if you have a protein with a disulfide bond ~ 40kda dimer, then to get mass as it travels it would be 80kda?
Under reducing, a homodimer of 25kda and a homodimer WITH disulfide bonds composed of 40kda substituents would ALL travel separately so you’d just see 25 and 40 monomers traveling down the gel.
Is the correct? Thanks!
They’re on standbi
Pilot on me!!
Nothing, he was gladiator.
Dad jokes are supposed to be jokes you can tell a kid and they will understand it and find it funny.
This sub is mostly just NSFW puns now.
If it needs a NSFW tag it's not a dad joke. There should just be a NSFW puns subreddit for that.
Edit* I'm not replying any longer and turning off notifications but to all those that say "no one cares", there sure are a lot of you arguing about it. Maybe I'm wrong but you people don't need to be rude about it. If you really don't care, don't comment.
When I got home, they were still there.
What did 0 say to 8 ?
" Nice Belt "
So What did 3 say to 8 ?
" Hey, you two stop making out "
The heme‐copper oxidase superfamily comprises cytochrome c and ubiquinol oxidases. These enzymes catalyze the transfer of electrons from different electron donors onto molecular oxygen. A B‐family cytochrome c oxidase from the hyperthermophilic bacterium Aquifex aeolicus was discovered previously to be able to use both cytochrome c and naphthoquinol as electron donors. Its molecular mechanism as well as the evolutionary significance are yet unknown. Here we solved its 3.4 Å resolution electron cryo‐microscopic structure and discovered a novel dimeric structure mediated by subunit I (CoxA2) that would be essential for naphthoquinol binding and oxidation. The unique structural features in both proton and oxygen pathways suggest an evolutionary adaptation of this oxidase to its hyperthermophilic environment. Our results add a new conceptual understanding of structural variation of cytochrome c oxidases in different species.
https://ift.tt/3uVQbXs
KRAS homo‐dimerization has been implicated in the activation of RAF kinases, however, the mechanism and structural basis remain elusive. We developed a system to study KRAS dimerization on nanodiscs using paramagnetic relaxation enhancement (PRE) NMR, and determined distinct structures of membrane‐anchored KRAS dimers in the active GTP‐ and inactive GDP‐loaded states. Both dimerize through an α4‐α5 interface, but the relative orientation of the protomers and their contacts differ substantially. Dimerization of KRAS‐GTP, stabilized by electrostatic interactions between R135 and E168, favours an orientation on the membrane that promotes accessibility of the effector‐binding site. Remarkably, ‘cross’‐dimerization between GTP‐ and GDP‐bound KRAS molecules is unfavorable. These models provide a vital platform to elucidate the structural basis of RAF activation by RAS and to develop newer inhibitors that can disrupt the KRAS dimerization. The methodology developed to specifically probe the intermolecular interactions within KRAS dimer is applicable to many other farnesylated small GTPases.
https://ift.tt/39vJYFT
