Planning Next

Sunday, January 19

162.4 lbs (Day 23, starting from 162.6 lbs on Dec 28)

Today is so simple to explain haha. Nothing special. I hit the gym in the morning and did the rest of the assignments from the reaction mechanism class. To be honest, I go between YouTube videos and assignments back and forth🫠 There is no way to escape from watching Pinggyego, which I mentioned several times… I managed to finish the assignment around dinner(I ate a delicious cioppino from Costco. again, I highly recommend it), wrote a blog post in some amount, and went to bed early for the next morning’s gym session!

Monday, January 20

161.6 lbs (Day 24, starting from 162.6 lbs on Dec 28) | 🏃🏻 Done (1/6)

Today was technically a national holiday, but I had work to do, so I headed to the lab. It’s not mandatory to go in on weekends(as well as on weekdays), but I like to go and get some things done. In my opinion, making weekend lab work mandatory is not a good long-term strategy. Once I arrived, I met my mentor, and we discussed the sequencing data we received yesterday from a vendor. Fortunately, one out of the four plasmids we made contained the mutation we wanted—though it wasn’t a perfect match for another non-critical part. It did have the desired amino acid mutation, but it also had several insignificant mutations elsewhere in the plasmid. We decided to make more plasmid by transforming our desired one into bacteria, just as we did before. About a week later (next Monday), we plan to express our target protein, purify it, and then do a labeling experiment with the warheads I’ve synthesized.

I saw a parrot(?) when I arrived home to have lunch. The bird seems not to be afraid of me haha. I pulled out my phone and took a picture and video, but as you can see in the above video, it was funny that the bird flew directly to my face. I was freakin out(See the last frame of the video, you can see)😳

Heading back to the work, I ran a nitro reduction using a nickel catalyst, ammonia (7 M in methanol), and hydrogen to convert the nitro group into an amine. I had done a similar reaction the day before, and it was successful. I also checked two NMR spectra. The first was for the amine salt I made yesterday from the free amine, and the other was (R)-acrylamide-type warhead. Because the initial reaction went well, I repeated it on the remaining reagents I had, this time on a larger scale.

I also tested DCM (dichloromethane) and THF (tetrahydrofuran) as solvents in another warhead synthesis reaction—specifically for an ethane sulfonyl fluoride compound. Acidic conditions with a nucleophile can cause decomposition of the fluoride, so I wanted to use a normal-phase column with hexane and ethyl acetate. For that reason, I planned to evaporate the reaction mixture and then load it directly onto the column. After letting the reaction run for about an hour and a half, I checked the LC-MS of the two reaction mixtures. The DCM-based mixture showed fewer side products and contained more of the desired compound, so I decided to proceed purification of this.

I knew the scaffold I was working with had a high affinity for silica, so I was worried about purification efficiency. My goal, though, was just to get enough of the desired compound for a labeling experiment, so I wasn’t too concerned about the yield. I started with a gradient of up to 100% ethyl acetate (over about 20–25 column volumes), but I saw a lot of tailing on the column. I switched to a DCM/methanol system, hoping that using methanol without acid additives (like TFA) wouldn’t facilitate the hydrolysis of the sulfonyl fluoride. Unfortunately, even after running another 20 column volumes (up to 15% methanol in DCM), I only saw a weak signal for the product in the LC-MS. That told me normal phase wasn’t ideal for purifying this type of scaffold.

Next time, I plan to try DMF, which has worked better in the past and produced fewer side products. I’ll also try using water and acetonitrile as eluents without TFA additives. This is because acidic conditions can protonate the sulfonyl group, increase the sulfur’s electrophilicity, and lead to hydrolysis of the sulfonyl fluoride. That’s my second strategy, and I’m going to give it a shot tomorrow.

After finishing the lab, I headed directly to the gym to work on my chest, triceps, shoulders, and abs. It was a delicious and good workout, but I was so exhausted🥱 When I got home, I ate dinner and literally passed out. I woke up around 12:30 or 12:40 a.m. and realized I still had laundry, meal prep, and some English study to do. Most importantly, I needed to prepare a conversational script for my meeting with a professor about my third rotation. I finally went back to bed at around 3 a.m.

Tuesday, January 21

162.8 lbs (Day 25, starting from 162.6 lbs on Dec 28) | 🏃🏻 Missed (1/6)

Woke up at 8:00 a.m., had breakfast, and headed to the lab. I worked up the larger-scale reduction reaction, then crystallized the product by adding HCl in dioxane. After filtering and washing with ethyl ether, I obtained a solid with a significantly improved yield. Another experiment I did today involved re-synthesizing the sulfonyl fluoride. Last time, the compound was hydrolyzed under reverse-phase chromatography with TFA in acidic conditions, so I tried normal-phase chromatography using ethyl acetate/hexanes and also dichloromethane/methanol. Unfortunately, I failed with both attempts because the product peaks were so broad that I couldn’t identify or separate the main product from the side products.

This time, I went back to reverse-phase chromatography but tried it without TFA. However, the product never eluted. In the end, I added a small amount of TFA (0.1%) in the acetonitrile line only, while keeping the water line free of TFA. I was hoping to reduce the overall acidity(dilution) to prevent degradation. Despite that, when I checked the sample by LC-MS, it looked like the product might have hydrolyzed anyway. I decided to confirm a side product I got by NMR later. I tried to lyophilize the sample, but the lyophilizer was down for maintenance, so I stored the samples at –20°C for now. That wrapped up my experiments for the day because I had a lot of things to do besides experiments.

Aside from lab work, I attended a QBC journal club class from 10:45 a.m. to noon. After lunch, I spoke with a professor whose lab I hope to rotate in during the spring quarter. However, because the outcome of a grant application won’t be known until after the spring rotation period, it was unlikely to ensure joining possibility after spring rotation. Our options were, first of all, after checking the grant results, doing summer rotation. Second, we need to find another co-advisor to help resolve funding uncertainties. We discussed the possibility of co-advising with the professor whose lab I am now rotating in. Unfortunately, it turned out not to be feasible because both professors’ research areas overlap significantly, compromising the purpose of co-advising, which is usually to combine two distinct areas of expertise. So, this arrangement wouldn’t meet that goal. The professor said there was still a possibility of finding another co-mentor. Still, I was unsure about that because I vividly realized the difficulty of being co-advised. I should have a reason for co-advising and a solid thesis project that both professors can give advice and are willing to develop.

Later, after work, I met my current rotation PI and discussed the possibility of joining the lab. It was not a 100% obvious expression, but the professor mentioned he already has two students from the same program who likely want to join the lab. The professor wasn’t sure if he could take a third student from the same program due to a program policy, so we concluded trying to figure out that there could be an exception to that policy. I said I would contact our program director to determine whether three students in one lab are allowed in a year. Suddenly, it hit me that joining the two labs I had been interested in might not be so easy, and that realization kind of made my head spin😵 I do think there’s still a chance for me to compete in my current lab. But even though joining a lab is not first come, first served, I’m not thrilled about squeezing my way in and making things awkward. Now I feel a ton of pressure to find another lab for my third rotation where I’m sure I can get in, and it’s pretty stressful.

In the evening, I went to Costco with friends who mentioned that the lean turkey at Costco has a distinct smell, which she really dislikes. Until then, I hadn’t noticed it, but after she pointed it out, I realized there was indeed a noticeable unpleasant smell😐 I decided to switch to beef for my meal prep, so I bought a large package of beef and made several meals with it. Delicious! Meanwhile, the professor forwarded me some interesting modeling data from Scripps. It described the binding sites for a small molecule we’re studying. He asked for my thoughts on their proposed model, so I spent around three hours researching, creating some initial models of my own, and reading background literature before emailing him my feedback. Time flew by because I really enjoy this kind of work!

Wednesday, January 22

162.0 lbs (Day 26, starting from 162.6 lbs on Dec 28) | 🏃🏻 Done (2/6)

From now on, I decided to organize things I have done each day into two parts: Outside of Work and Lab Work. I hope this helps both me and my readers follow the flow of my day easily.

Outside of Work

I woke up at 6:00 a.m., went to the gym, and had a good workout. Unlike the typical winter seasons in Korea, these days I have very nice weather, which makes me take pictures of the clear sky. I had a class from 1:00 p.m. to 2:30 p.m. with a different professor than last week. He was very engaging and tried hard to involve the audience. The topic was oxidation chemistry, and we learned about several new reagents I hadn’t encountered before(too many…).

After class, I talked with another student rotating in the same lab. We discussed the possibility of joining this lab and the current situation. Many people are interested in joining, and when I spoke with the professor yesterday, it sounded like he might accept two students this year due to the program’s policy. Because of this, we plan to contact our program director to explore our options. Typically, a single PI can take a maximum of two students each year, but there are exceptions—sometimes a professor can take “2.5” students if one is co-advised with another lab. We don’t want to compete with the other candidates, but we also want a fair chance at joining a lab we’re passionate about. We hope meeting the director will help us figure out a solution.

Lab Work

Yesterday evening, the professor emailed my mentor and me some interesting protein-binding site modeling data for one of our designed compounds. We’re studying the rationale behind the proposed binding mode, so my mentor and I discussed the details of the molecule and the binding pose suggested by the model.

On the synthesis side, I tried again to purify a sulfonyl fluoride compound. I asked a postdoc who had been working on similar compounds for the current synthesis problem. He explained the chemistry of sulfonyl fluoride really well, which prompted me to try a normal-phase column with 2% acetic acid as an additive. When I ran the TLC, I saw two main spots: the top spot likely represented the dialkylated product, and the bottom spot presumably the monoalkylated product we wanted. However, there was some trailing on the plate. After purification, I checked the fractions by LC-MS, but the desired M+H ion for the monoalkylated product was either absent or too low to detect. I plan to synthesize other warheads starting tomorrow.

Finally, I ran a Chai-1 prediction on our protein of interest and the designed compound. The modeling data from Scripps (produced by AutoDock, a physics-based model) is interesting, but I wanted to compare those results with a more AI-driven method. Surprisingly, the model successfully (even more accurately) predicted the preference of ligand positioning in the binding site. I don’t know how accurate the Chai-1 prediction is, but I forwarded the results to the professor and my mentor.

Thursday, January 23

160.8 lbs (Day 27, starting from 162.6 lbs on Dec 28) | 🏃🏻 Done (3/6)

Outside of Work
Woke up around 6:00 a.m., and I freaked out that my weight hit the lowest one within the last month. It might be because I am just too worried about my future (thesis lab), so that case I kind of easily lose my appetite and become not diligent in eating food and water. I don’t think it is serious, considering not drinking water surely makes body weight lower fastly. I will manage more thoroughly. I managed to get to the gym for a lower-body workout. It’s always tough, but it’s important not to skip it if you want to maintain balance between your upper and lower body. In the gym, I came across a graduate student in the lab where I first rotated. We discussed the possibility of using AI for small-molecule drug discovery, specifically how to use Chai-1 on the UCSF computational cluster because now I am also considering co-advising.

I attended the BP/CCB Pizza Talk from 5:30 to 7:00 p.m. It had been a while since I last went, but I decided to join because a professor whose work I’m interested in was presenting. Surprisingly, the more interesting research talk was given by another professor who had literally just begun as a PI this January. If the professor defines the interactome near the DNA-binding region more concretely, it will be critical data for any other researcher interested in studying transcription. I thought this was so impactful research. That presentation made me contemplate how I can maximize my influence while still pursuing the research I want to do in my PhD. After the pizza talk, I asked one of my cohort members, who had already decided to join a synthesis-heavy lab, why he chose that path. We had a deep conversation about the meaning of a PhD, their worries, and their beliefs. Many friends shared their opinions on the topic, and it was really helpful for me. Thanks, everyone!

Lab Work

This morning, I realized there was a problem with the chirality of one of the molecules we’ve been studying. I had submitted both the (S)- and (R)- forms of the ligand as SMILES to Chai-1, but the results always came back in the (S)-form—even when I specifically submitted the (R)-form. When I checked the docking poses generated by AutoDock Vina from Scripps, I noticed they, too, were all in the (S)-form. This is concerning because our cell-based assays and other biological experiments show that the (R)-form of our small molecule is more potent. In response, my mentor emailed our collaborators to clarify the issue and decide on the next steps.

For the past three or four days, I’ve also been working on synthesizing a sulfonyl fluoride–type warhead. We decided to postpone it for now because purifying the intact product has been difficult. Although LC-MS indicated some products immediately after the reaction, the purification process turned out to be problematic. I looked into various coupling reagents, including PyBOP and HATU, and also recommended trying COMU for the synthesis of other warheads.

Friday, January 24

161.4 lbs (Day 28, starting from 162.6 lbs on Dec 28) | 🏃🏻 Done (4/6)

Outside of Work
Afternoon, Reaction mech. class as always. In the evening, I went to the gym, and I was so exhausted that I literally passed out afterward. Lately, I’ve been feeling a lot of uncertainty about my future, along with a heavy workload. Adding to this stress, there has been no response from the director regarding our discussion about the policy limiting the number of students who can join under one PI. I really hope things will start to move forward soon!

Lab Work
I had a quick meeting today about the project I’m working on, but from a different angle: exploring possible binding poses of our ligand with the protein of interest. I had prepared slides to update everyone on my progress, thinking I might present. But, I didn’t get the chance since a former postdoc, who had previously sent us some modeling data, joined and had updates to share. So there wasn’t much time left. However, I shared my Chai-1 prediction and interpretation, which sparked a great discussion with plenty of feedback. It was a valuable experience I’d never had before. Everyone, including the current PI, seems to have a solid understanding of what’s going on with the project, making the meeting worth attending.

Not much on the experimental side today: I just checked the LC-MS results for three different coupling conditions from yesterday, then quickly purified two compounds, which went smoothly (based on the LC-MS patterns😄). After that, I proceeded with lyophilization. That’s all~

Saturday, January 25

163.0 lbs (Day 29, starting from 162.6 lbs on Dec 28) | 🏃🏻 Done (5/6)

Yesterday, I went to sleep at 8 PM, so I woke up at 3 AM, did the house chores, and went back to bed at 5 AM😵 It was my grandmother’s birthday, so I made a short call. Being away from my family is the hardest thing about studying abroad. Especially considering my grandmother’s age, I want to finish my degree as soon as possible and go back to Korea. Of course, I have dreams and a future that I want to pursue, but sometimes I feel like I have given up so many things for them. Every time I think about this kind of stuff, I wish for my family’s well-being and health. The most evident thing I must do now is focus on my studies, complete my degree well, and maintain a healthier mental and physical state. I also hope to invite my whole family to my thesis defense!

Due to the Reaction Mechanisms class, our cohort started a study session for the class two weeks ago. At that time, I had appointments, so I was unable to attend. Today, I had no events to go to and had to do my assignments, so I headed to the room borrowed by some of my cohort members. On the way, I bought a heavy(so heavy) burrito filled with three kinds of meat from the taco truck in front of my home. The taste and amount matched the cost 👍 To be honest, I couldn’t finish it on the spot and ended up eating it for about an hour. + Today was the birthday of one of my cohort members. He brought delicious cupcakes for us. Thank you very much🥳

After meeting my friends, we discussed many questions to which we had different answers and details. I realized again that my English still has a long way to go. I don’t have much experience explaining basic chemistry principles in English, so when I had to use them to explain organic chemistry concepts, it was really tough. But this session was much better than times when I had to explain things I wasn’t sure about. When I have confidence in the content and my logic, it’s relatively easier to lead and maintain the conversation. It gives some breathing room to focus on my grammar and English itself, not just the content.

After the study session, I headed straight to the gym. I was happy that even though this week was kind of busy and chaotic (because I was struggling with thoughts about my future path), I still managed to get five gym sessions💪 Right after a quick meal, I went to the Hub to finish the rest of my assignments. You guys might remember that I wasn’t worried about the class, but this week in lectures, there were so many reactions I had never studied before. To be honest, I didn’t like the class structure, where they just kept listing as many different organic reactions as possible. There must be a reason why many researchers have developed new methodologies to drive the same transformations using different reagents. Of course, if I take extra time outside of class to study various reactions individually, I would understand the logic, but it feels like overkill haha. It feels like studying never ends, no matter how much I do.

After almost finishing my assignment, I tried to narrow down my third rotation option once again. I decided on my last (hopefully🙏) rotation lab. I already heard that the professor wants to have rotation students and new graduate students, so I don’t think there’s a possibility that I won’t be able to join the lab after the rotation (of course, I will try my best 🏃🏻). I am a little worried, though, because even though the lab publishes in top journals at least once a year with relatively few people (<8 members, including postdocs), there have been many stories about the lab from my cohort as well as the senior cohort. But after talking with a graduate student in the lab, I felt it wasn’t like that, and I thought it was worth trying. One good thing is that the research in this lab is similar to what I am doing in my current rotation lab(just methodologically). Anyway, I wrote an email to the professor about the possibility of a third rotation and am now waiting for a response!

조금 늦었지만 다들 새해 복 많이 받으세요!🧧🎊