This came up as I was skimming the internet…
Jimmy Kimmel asked people on a gluten free diet: “Do you know what gluten is?”
Hahaha, you can guess the answers….
http://www.huffingtonpost.com/2014/05/06/gluten-free-people-have-no-idea-what-gluten-is_n_5273980.html?utm_hp_ref=mostpopular
So today was our last day of orgo lab, and we all presented on our drug synthesis papers. In my paper and in my presentation today, I explained the synthesis and biochemical function of azithromycin, also known as Zithromax or Z-Pak. This molecule is a 15-membered macrolide antibiotic. A macrolide antibiotic is a molecule/drug containinga macrocylic lactone ring and 2 attached sugars. Macrolide antibiotics come in many shapes and sizes, but what is unique about azithromycin is its extended half-life compared to other macrolides. While most other macrolides have either 13 or 14 membered macrocyclic lactone rings, azithromycin has a 15 membered ring. This difference, and several other differences, allow for azithromycin to have about a 11-14 hour half-life, compared to say, erythromycin A, a 14-membered macrolide, that has about a 6 hour half-life. Because of this long half-life, azithromycin is actually in a much higher concentration in your macrophages/tissues than other antibiotics when you take it, which allows doctors to be able to prescribe it over a much shorter amount of time than other antibiotics, usually only for 5 days. (Some antibiotics have to be taken for 21 days to complete their effect!) Some causes for concern lately, however, involve studies that show that azithromycin may actually increase the QT interval during heart muscle depolarization. Basically, the QT interval measures the length of a single heart beat, so they are seeing that in some patients who take azithromycin are developing these uneven heart beats, also known as arrhythmias. And I had heard this from my dad who works with a lot of these drugs/the news, but after more research, I found that these studies only claim that azithromycin has about a 0.8% increase in arrhythmias compared to other antibiotics, and that it was really only seen in patients with pre-existing heart conditions.
Anyway, what I was interested in was how it functions to combat bacterial infections. And it turns out, its super cool! So as you may know from your vast knowledge of biology, ribosomes have two main subunits, a larger one that sits on top of the mRNA, and a lower one that essentially caps the bottom of the mRNA to the rest of the ribosome. In bacteria, the larger subunit is called the “50S” subunit and the smaller one is called the “30S” subunit. So as we know, ribosomes zip across the mRNA, synthesizing a growing peptide chain out from this top, larger subunit. So what I found is that azithromycin actually squeezes itself into the center of this 50S subunit in bacteria, and blocks the addition of any amino acids to the growing peptide chain, arresting any protein synthesis in bacteria. And without being able to produce any proteins, the bacteria die! Below is a PyMol figure I made showing the 50S subunit, with azithromycin in dark blue in the core of the protein. Actually, inhibiting protein synthesis by basically ‘plugging’ the 50S subunit of bacterial ribosomes is what all macrolide antibiotics do, being just one subsection of many types of antibiotics.
Also, during my paper, I started to think about just how crucial it is for azithromycin and these macrolide drugs to interact selectively with bacterial ribosomes, since if they ever interacted with ours too, we would be not so happy (also dead). However, human ribosomes are much larger than bacterial ribosomes (we have 60S and 40S subunits) and are shaped somewhat differently. I believe this may either cause our ribosomes to have differently shaped binding sites and that azithromycin may not have a compatible shape to insert itself there. Also, its possible that the polarity may be slightly different in human vs. bacterial ribosomal binding sites, which would make azithromycin have more favorable interactions with bacterial ribosomes (as you can see, it has many hydrogen bonding possibilities, as well as defined hydrophobic sections). Below I show both the macrolide structure of azithromycin, and where it inserts itself into the 50S subunit. Pretty cool, right!
I just listened to this little story on NPR about biochemists at Scripps Research Institute in La Jolla, California (across the street from my high school!!) who are attempting to create two new synthetic bases for DNA. Honestly, my gut reaction is that this is kind of a biochemical blasphemy, haha. They say, ‘just imagine what we could do with infinitely more genetic code combinations, and imagine being able to code for so many infinitely more things in our bodies’. This actually kind of scared me. I feel like the system we have now is what we have because it functions so seamlessly together (demonstrated by all these presentations we’ve been having too) and is incredibly efficient and diverse despite having only 4 bases. So many more changes would have to be made for us to even process these new bases. DNA polymerase, DNA’s shape, enzymes that repair DNA, RNA polymerase, RNA bases, size of histones, size of chromosomes, transcription factors – all of these and infinitely more tools for processing out genetic code in the cell would have to be changed. Not to mention if our cells mess up while reading these new bases with these new tools, cancer would quickly become an even larger problem. Not sure how I feel about it!
http://www.npr.org/blogs/health/2014/05/07/310282870/chemists-expand-natures-genetic-alphabet
I recently came across this interesting condition called dermatographic urticaria, also known as dermatographism or “skin writing”. The more common name for urticaria is hives, or bumpy/red skin rash that may be caused by allergies. Dermatographism is a prevalent type of urticaria, especially in young adults. If you have a friend with “sensitive skin” that reddens and swells upon contact, s/he may have actually have dermatographic urticaria.
Most interestingly, this is thought to be caused by the release of histamine from mast cells, which are involved in the allergy pathway. The membrane around the mast cell is weak and breaks under physical pressure, which causes the characteristic raised red skin.
http://www.ncbi.nlm.nih.gov/pubmed/6166647
Since we’ve been studying lipids, I though this was an appropriate reminder of how important the cell membrane is. The composition of the phospholipid bilayer directly affects its fluidity, and in this case, it is too fluid. The cholesterol, protein and lipid concentrations may need to be adjusted, with more long-chain saturated phospholipids.
We mentioned in class some time ago that micelles could be potential drug carriers in their aqueous core. Since my research from the past two years has been on drug and vaccine delivery (“controlled-release”), I perked up a bit at this concept, and thought I might share with you my research. I make micro- and nanoparticles out of polymers using a water-oil-water double emulsion. In my current project, this means I mix a small amount of water (plus iron oxide particles and fluorescent dye) in with polymer (PLGA-PEG) dissolved in an organic phase (DCM). The water phase “hides” from the organic phase by clumping together into little pseudo-particles. Then, I drop this mixture into an outer aqueous phase, PVA, which forms “bubbles” of polymer due to the hydrophobic effect. However, if I was good about homogenizing the first mixture, the polymer will encapsulate most of the water with the Fe2O3 and dye in aqueous phase inside the polymer shell. The resulting particles are thus water (PVA) on the outside, oil (polymer) for the shell, and water (Fe2O3 and dye) on the inside. The same principles apply to micelles, if we replaced our synthetic organic phase with a natural lipid.
In Week 4, we also talked about biological symmetries. Didem mentioned that the poliovirus has icosahedral symmetry. This past fall, I actually spent a fair amount of time testing the efficiency of self-assembly and strength of different shapes for a gastrointestinal “pill” that would self-assemble in the gut and get caught in the intestines to slowly release drug as the polymer delivery system degraded over the course of a month or more. In reality, I glued magnets to a lot of plastic shapes and flipped them around together in a cup. Living the dream! I can’t share my results here (or else we might not get the patent!), but we did find that cubes, dodecahedrons and icosahedrons are all more stable than tetrahedrons and octahedrons. So, yes, I have scientific evidence that the poliovirus is efficiently built!
Also, I hope that you enjoyed my pun!
We’ve been spending these last weeks on cell membranes, but new research has discovered the origin of life before membranes. Life before cells, you ask? That’s exactly right. Recent findings suggests that there are such things as metabolisms without bodies. A graduate student at the University of Cambridge found, in his new media, pyruvate — the product of glycolysis — but there were no cells in his media to have produced this compound. He and his PI rushed over to the Earth Sciences department, where they confirmed that such metabolic processes, which produce ATP and are vital to all life on Earth, are indeed capable of operating independently from cells and our lipid membranes in the hot, metallic waters of Earth 4 billion years ago.
As we close this year, I thought that this finding nicely reminds us of what we’re really studying here. Following the evolution of life from the loose metabolic systems by the thermal vents of young Earth to the complex individuals we are today — with our complex emotions, symbiotic microbiota, and entrepreneurial capabilities — we truly appreciate how the biological processes we are studying grow up in this world. Starting with a free metabolic system, we can understand why lipid membranes would be more successful. In this amazing TED talk, Martin Hanczyc shows protocells whose lipid membranes (oil spots) trap simple, 5-chemical compound metabolisms to enable his protocells to move and even replicate (surprise!).
With the evolution of life, we are getting to see this amazing interplay developing between form and function, and between function and form. We see RNA, perhaps catalyzed by inorganic clay molecules, replicate before protein polymerases developed to assume this role more efficiently, membranes entrap metabolisms, and RNA introns (miRNA) take over regulatory roles. As we zoom into the present, we wonder at the intricateness of carbohydrate antigens marked by our immune systems and nucleic acids controlling whole organisms. Biochemistry — It’s Amazing!
Sources (from Prof. Arumainayagam’s CHEM 306):
http://www.newscientist.com/article/dn25471-spark-of-life-metabolism-app#.U2OrB61dXAg
http://www.ted.com/talks/martin_hanczyc_the_line_between_life_and_not_life
In the chaos of this last week of classes, it is easy to get lost in exams and the mucky details of classes. So I was happily surprised today when, during final presentations for my Organic Chemistry class, I had a minor epiphany and realized how much we’ve learned since the beginning of the semester.
In orgo, we were presenting on compounds that we researched throughout the semester, most of which had some biological relevance. First of all, one person had a compound that interacted with glutathione (which to most of you should sound a bit familiar…). Another poster also displayed some of the interactions between the compound and a protein receptor, and I could easily identify the residues involved and what types of reactions were going on just from the small picture!
Its hard to remember what we didn’t know just a few months ago, but I’m really glad I got the chance. If any of you have a second to think about it, its really rewarding!
After spending the last few weeks of lab with fluorescence spectroscopy, I was reminded of an equipment I have used previously in the lab I used to work in, called flow cytometry. This piece of equipment can be used for cell counting, cell sorting, and biomarker detection. The beauty of it lies in its ability to simultaneously provide multi parametric analysis very quickly. I used this machine in the context of trying to identify surface markers that are commonly found in leukemic cells. So how exactly does flow cytometry work its wonders?
It turns out that flow cytometry is based on fluorescence. For example, one has to use fluorophores as labels to attach to an antibody that recognizes a target feature of the cell. Each fluorophore has a characteristic peak of excitation and emission wavelength. The fluorophores attached to cells are excited by laser and the emission spectra is collected, and using such data, a very specific set of cells can be gated and analyzed because of the specific flurophores that had been used.
Often when you look at the ingredients list for lotions, sodium hydroxide is listed way down on the bottom, and they quite commonly found (ex. Nivea).
So what exactly is such a strong base used for? When we made soap, we also used it. Just a recap of why we needed the strong base, the oils that we started off with to make the soap were fatty acids, which are quite hydrophobic. The NaOH makes these fatty acids into salts of that fatty acids. This is why a soap works – the Na+ end can dissolve in water and the carbon chain can dissolve in the oils, and this makes it possible to wash away the oil using water.
For lotion, the NaOH is used as a neutralizer in a very low concentration that would not cause sever burning. NaOH neutralizes the acidic fatty acids, like stearic acid which is another in credit, to make the carbon chains more friendly to our skin.
The lab I’m a part of does a lot of work with anti microbial peptides whose mechanism of action is either permeabilizing the membrane (effectively poking holes) of the target cells or translocating through the membrane into the cell and into the nucleus to interact with DNA. I’ve wondered how safe these peptides would be for use considering that both bacteria and human cells have membranes but understanding the composition of the membranes of the two kinds of cells has explained why. It turns out that though most bacterial membranes are composed of neutral phosphatidylethanolamine, they also contain many liposaccharides which give them a net negative charge. Human cell membrane contain varied lipids and charge is not negative like the bacteria’s. As such, the polycationic anti microbial peptides can target the negatively charged bacterial membranes but leave the human ones intact.