Imagine protein X, destined to span the plasma membrane; assume the mRNA carrying the genetic me

step 1 Hello there, students. Today we're going to be discussing the question that is right in front of

Imagine protein X, destined to span the plasma membrane;...

Key Concepts

Rough Endoplasmic Reticulum (ER)

The rough ER, characterized by ribosomes attached to its surface, is the cellular site where membrane-bound and secretory proteins are synthesized and translocated. For a protein destined to span the plasma membrane, the rough ER provides the machinery needed for the proper insertion into lipid bilayers, folding, and initial post-translational modifications such as glycosylation. In cell fractionation experiments, the rough ER and its associated proteins, including those being processed for the plasma membrane, appear in the microsomal fraction.

Vesicular Transport and Protein Trafficking

After initial synthesis and membrane insertion in the rough ER, proteins destined for the plasma membrane are packaged into transport vesicles. These vesicles bud off from the ER and move through the Golgi apparatus, undergoing further modifications, before being transported to the plasma membrane. This vesicular trafficking pathway is essential for ensuring that proteins reach their appropriate location on the cell surface where they can perform their functions.

Co-translational Translocation

This concept refers to the process where a nascent polypeptide is inserted into the endoplasmic reticulum (ER) membrane during its synthesis. As the protein is synthesized, a signal peptide emerging from the ribosome directs the entire complex to the ER, where translation continues into or through the ER membrane. This mechanism ensures that proteins, especially those destined for the cell surface or secretion, enter the correct cellular compartment immediately upon synthesis.

Signal Peptide and Signal Recognition Particle (SRP)

A signal peptide is a short amino acid sequence at the beginning of the newly forming polypeptide that acts as an address label for the protein, directing it to the ER. The SRP, a ribonucleoprotein complex, recognizes this signal peptide as it emerges from the ribosome, temporarily halting translation and targeting the ribosome-mRNA complex to the ER membrane. Once associated with the ER, translation resumes, allowing the protein to be co-translationally inserted into or across the ER membrane.

Transcript

00:01 Hello there, students.

00:02 Today we're going to be discussing the question that is right in front of us.

00:06 So the question reads, imagine protein x was destined to span the plasma membrane.

00:14 Let's assume that mrna was carrying a genetic message for protein x, where it has to be translated by ribosomes in a cell culture.

00:25 If the cell was fractionated, which part of the fractionation would protein x be found? explain this by describing this transit.

00:34 And obviously down here, you could tell this is the process to do a cell fractionation.

00:41 And for those who don't know, it's just basically a process of taking the cells apart and separating its major organelles and other subcellular structure from one another.

00:55 To basically get the desired components out of the cell so we can learn more about the cell that is right in front of us.

01:05 So the process starts off with this test tube that has cell tissues, i mean tissue cells, and it's put in a blender where it's going through homogenization, which is basically blending into one.

01:21 And the next test tube we see that it's been homogenated, then they put it through a process called centrifification, centrifugation, my bag, which is basically you're putting the homogenated cell tissue in a machine where it's spinning the test tube at a high speed, for a certain amount of time between seconds, minutes, hours, to even for some days.

02:01 I know it's crazy, but there's some who have, or i just know someone who's crazy enough to do it for days, but, or a whole day.

02:11 So then this side of the test tube has been centrifuge.

02:15 And i wrote on the top a thousand times the force of gravity for 10 minutes.

02:21 That's why you see down here is a thousand, thousand grams because that's how much of the pellet that was removed from the cell.

02:35 So let's look at the first test tube.

02:38 The first test tube, so with the first test tube, you can see the separation between a supernatic.

02:48 Supernatic.

02:50 And supernatic is just basically the liquid that's above the pellet.

02:53 And the pellet is where it has a desirable components of your major organelles and other subcellular structure that you're looking for to learn about the cell.

03:04 So for this first pellet, it's rich in nuclei in cellular debris.

03:14 So then after that, you want to separate and go further down, you will pour the supernatid component into.

03:25 The next test tube and this one spend it for they end up being centrifuge for 20 minutes which they end up getting 20 ,000 grams.

03:36 And the palette down here is rich in mitochondria.

03:42 And i also put chlorplas down there because that would work if you're studying the plant cells.

03:53 But since we're talking about animal cells, we're going to focus on this for the time being of the discussion.

04:01 And then you do the process over again, and this time they did it for 60 minutes.

04:08 Yeah, basically they did a whole hour where they got 80 ,000 grams, where they end up getting a pallid, a pellet that's rich in microsomes.

04:20 The microsomes, it's just pieces of plasma membranes and cells internal membranes.

04:28 So you're getting all that within the cell membrane.

04:32 And then going over one more time where you end up getting 1 ,500 ,000 grams, and they spend it for three hours where the palate, i mean, the pellet is rich in ribosomes.

04:48 And with this process, you see, is basically called differential centrifugation...

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