Episode 26 is our first listener’s choice episode! The first recAP is about membrane proteins - both structure and function.
Episode 26 is our first listener’s choice episode! The first recAP is about membrane proteins - both structure and function (1:18). Second, a biotechnology refresher with discussion of bacterial transformation, electrophoresis, and PCR (3:10). Lastly, Melanie reviews each scientist’s contribution to the discovery of DNA as the hereditary molecule (5:51).
The Question of the Day asks (7:40) “What percentage of DNA is made of nitrogen?”
Thank you for listening to The APsolute RecAP: Biology Edition!
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Hi and welcome to the APsolute Recap: Biology Edition. Today’s episode will recap multiple topics - it’ our first Listener’s Choice Episode.
Lets Zoom out:
We all have biology topics we feel more confident in than others. We’ve also all likely lied to ourselves on occasion about what we know and understand. Does this sound familiar to you? You have a stack of flashcards and look at the first one. Before even giving your brain a moment to think, you flip it over to read the answer going “oh yeah, I knew that.” Today’s episode will dive into some of the concepts we need a refresher on. We want to thank everyone who wrote in with topic requests. I’m sorry we couldn’t get to all of them - but we will have more listener’s choice episodes in the coming weeks! If you have a topic you’d like a recap on, please contact us through email, instagram, or twitter.
Lets Zoom in:
Our first recap topic comes to us from listener Davian - membrane proteins (with three exclamation points). Proteins are polymers (or polypeptides) of the monomer amino acids. An amino acid is made up of one central carbon atom that forms four single covalent bonds – one is to hydrogen, another to a carboxyl (c=o, o) and a third to an amino (NH3). The fourth covalent bond is to an “R” group. This R group is a substitute for different side chains that give each of the 20 different amino acids their unique properties. Proteins have four levels of structure as it bends and folds in upon itself. Primary is a chain of amino acids, secondary can either form an alpha helix or a beta pleated sheet. Tertiary is alpha helices and beta sheets folding further inwards. The final is quaternary, which has two separate amino acid chains, bent and folded and interacting together. The specific structure that each protein has is dependent upon the chemical properties of the R group.
Membrane proteins interact part way with the plasma membrane, completely cross the phospholipid membrane, or be loosely attached to its intracellular or extracellular components. Integral membrane proteins will have hydrophobic regions that interact with phospholipid tails and hydrophilic regions that are adjacent to the heads. Some integral proteins will have specific molecular chemistry internally, forming a channel. These channels may be gated or not and allow specific ions or small molecules to cross the membrane through facilitated or active transport. One of the most infamous integral carrier proteins is the sodium potassium pump. Peripheral proteins are more loosely attached to the membrane and are involved in cell recognition and communication. Examples include hormones and antigens.
Our second topic is biotechnology - requested by listener Jana. This is a biggie - but we will recap bacterial transformation, electrophoresis and PCR. Bacteria have a circular chromosome that can take in smaller plasmids. This process can be used in DNA cloning and you may have done this in class in the pGLO lab. Specific bacteria are mixed with DNA from a ligation and given a heat shock. This causes some of the bacteria to take up the plasmid. The bacteria is then plated on agar, sometimes treated with antibiotics or arabinose. After incubation, the bacteria which took in the plasmid will grow and express the new plasmid gene. In the case of the pGLO lab, the E.Coli expressed a glowing jellyfish gene.
Next, Electrophoresis. This is a process used to separate DNA fragments according to their size. A gel box has electrodes at either end and is filled with a salt buffer solution that conducts current. The DNA samples that were treated with specific restriction enzymes are pipetted into wells in a gel chamber nearest the negative electrode. Because DNA fragments are negatively charged due to their phosphate group, they migrate toward the positive electrode according to size. Small fragments of DNA containing fewer base pairs move farther down the gel than larger ones, containing more base pairs. Once the gel has been stained, you will see the DNA fragments as distinct bands. This process is often used to identify samples of DNA to a control group (as in crime scene samples to a suspect). You will need to measure and graph this data on a logarithmic scale.
Last for biotechnology is PCR or polymerase chain reaction. This process makes you a lot of copies of a fragment of DNA. This is useful in preparation of cloning experiments, or in forensics or medical diagnostics. You have a test tube of your small piece of DNA, a salt solution, primer, polymerase and free nucleotides. Denaturation separates the double helix at 96 degrees celsius. The test tube of your DNA sample is cooled to 55 degrees celsius for primer annealing (so that the primer will bind to each DNA strand). Then the sample is heated again to 72 degrees celsius for primer extension. With each heating and cooling cycle, the growth of DNA strands is exponential (1 strand becomes 2, 2 becomes 4 and so on). PCR that runs 20 times will create 1048576 DNA copies in just a few hours.
Our final topic for today is historical DNA Experiments - requested by Motnatty on Instagram. You don’t have to memorize their findings, but a background understanding of why and how the central dogma works is important. First, we had to figure out which was the heritable molecule and second, what was its structure? Griffith did his experiment injecting mice with bacteria. His transforming principle got so far as to say that something from the heat-killed bacteria “transformed” the rough bacteria and made them lethal. Avery took his work a step farther to determine what the transforming molecule was. Through chemical tests observing the ratios of nitrogen to phosphorus, Avery showed that the chemical composition of the molecule matched DNA, not protein. In addition, adding enzymes that break down DNA made the transforming principle inactive. The addition of enzymes that break down proteins had no effect. Hershey and Chase then came onto the scene with bacteriophages. A bacteriophage is a spacecraft shaped virus made of protein and DNA. They tagged the protein with radioactive Sulfur and the DNA with phosphorus. Results showed that only the radioactive phosphorus had entered the bacteria, again confirming that DNA and not protein was the molecule of heredity. We will recap Watson, Cricke and many others in our Scientists Recap with Episode 28.
To recap….
Membrane proteins are highly folded polypeptides that embed into the phospholipid bilayer for transport and communication. The three primary tools with biotechnology are bacterial transformation, electrophoresis and PCR. The battle for heredity involved multiple experiments to show that it is DNA and not proteins which contain the ingredients for life.
Today’s Question of the day is about DNA
Question. What percentage of DNA is made of nitrogen?