Mass Spectroscopy and Metabolomics

My name is Rob Parry, and I began my summer research a few weeks ago in Dr. Ian Blair’s Pharmacology Lab at the University of Pennsylvania.  The lab studies metabolism using mass spectroscopy techniques.  Mass spectroscopy is a fancy technique that detects molecules based on their mass to charge ratio by ionizing the sample (giving it a charge), exposing it to a uniform magnetic field, and detecting how much the magnetic field deflects the molecules flight path.  With this information, the mass spectrometer calculates the mass to charge ratio of the molecules it detects. Mass spectroscopy is a sensitive method to detect various metabolites and can accurately quantify how much of each metabolite there is.  My main project is to study the effects of inhibiting Coenzyme A (CoA) biosynthesis utilizing mass spectroscopy techniques.

Unknown-1 Coenzyme A

CoA is a molecule common to most, if not all, organisms and generally acts as an acyl-group carrier. CoA forms high energy thioester bonds with metabolites in a number of metabolic pathways including but not limited to the Krebs Cycle, fatty acid biosynthesis, fatty acid beta-oxidation, and isoprenoid biosynthesis.  It is estimated that CoA and its thioester derivatives take part in 4% of all cellular reactions in E. Coli (Wow!).

Pantothenate Kinase (PanK) is an enzyme crucial in CoA biosynthesis: it catalyzes the phosphorylation of pantothenic acid in the rate limiting step of CoA biosynthesis.  Genetic defects in PanK can result in neurodegeneration.  In addition, preliminary studies have shown that rotenone, a toxic molecule with a known association to Parkinson’s Disease, also impairs CoA biosynthesis.  Simply put, studying the inhibition of CoA biosynthesis may lead to insight on how neurodegenerative diseases come about.

In order to inhibit CoA biosynthesis, we are using hopantenic acid, a previously described competitive inhibitor of PanK.  Initial experiments were to confirm that hopantenic acid inhibitis CoA biosynthesis.

Unknown        pantothenic acid

Unknown   hopantenic acid

Isotopic labelling studies of pantothenic acid, a precursor of CoA and the natural substrate of PanK, were conducted.  If hopantenic acid affected CoA biosynthesis, we should have seen decreased incorporation of isotopically labelled pantothenic acid in Coenzyme A in cells exposed to hopantenic acid.  After 2 attempts, we saw did not see diminished labelling due to the presence of hopantenic acid (bummer).  Studies with longer incubation time and higher concentrations of the inhibitor in fresh media enriched with isotopically labelled pantothenic acid will be conducted next.  However, a few mishaps in cell culturing have caused delays in that project.

In the meantime, one of the other project we have been working on is “LC-MS analysis of human platelets as a platform for studying mitochondrial metabolism” for the Journal of Visualized Experiments.  The lab has previously used platelets as a platform to study the metabolic effects of various drugs.  Platelets are useful as they can be easily obtained in a large quantity from donors or from blood blanks.  While cancer cell lines are often used to study metabolic effects of various drugs, cancer cells metabolize differently than normal cells.  In addition, as platelets lack a nucleus, metabolic alterations in platelets are independent of transcriptional regulation, which can be useful to specifically study mitochondrial metabolism.  For these reasons, platelets can offer a useful platform for studying mitochondrial metabolism.

I am enjoying my time here and I am learning a lot both about metabolism and mass spectroscopy as a technique.  The people in the lab are all nice and very knowledgeable.  I get to run cool experiments, and, sometimes, I even get to play with dry ice.

Chemistry, Haverford, and Hydrogels

Three weeks ago, I started working in Dr. Karin Akerfeldt’s lab at Haverford College with my fellow lab mates Neal Patel, Ruben Aguilar, Vicky Isaac, and Marco Rivas. Cleaning and organizing the lab was the first task we set out to complete, and it became quite a bonding experience. Recently Neal, Ruben, and I (the resident rising seniors) made a mini scavenger hunt for Vicky and Marco (our rising sophomores) to help them become better acquainted with the lab. It was a tight contest, but Marco ended up being the winner!

Despite these fun activities, we are all hard at work on our respective projects. Neal, Vicky, and Marco are working on synthesizing (for the eventual crystallization) various calmodulin binding sites. Soon they get to head off to Sweden with Karin to continue the project with her collaborators! Ruben is focused on synthesizing gold nanoparticles of different sizes for eventual toxicity studies in Daphnia.

My project investigates hydrogels. Hydrogels are cross-linked peptide sequences that have the ability to absorb water. They are commonly used for drug delivery, cell culturing, and burn healing. My specific project investigates a peptide sequence from Semenogelin I (found in semen) that has been found to form a hydrogel in solution at neutral pH. To do this, I will be synthesizing, purifying, and characterizing various derivatives of this parent sequence with the hope to further understand of the gel formation of the parent sequence.

So far I have spent most of my time reading up on the history of this project and on the HPLC (high performance liquid chromatography, which is the how I’m purifying my sequence). My lengthy time of purification has not been without excitement however, for not only do I now have enough pure peptide for characterization studies, I formed my first gel last week (although by accident)! This was very surprising since the gel formed at an acidic pH rather than the neutral pH found to be needed for gel formation for the parent sequence. This week, I have been using Circular Dichroism and Fourier Transform Infrared (FT-IR) spectroscopy to start characterizing the gel formation of these sequences. Aka I get to make and play with more hydrogels!

Stay tuned for future posts on mine, and my fellow bloggers summer experiences!!!
– Keri Godbe ‘16

Aller guten Dinge sind drei*: German, Preposition Metaphors, and the Brain

—Claire Dinh ’16
*All good things come in threes.

WILLKOMMEN!
In German, that means, “Welcome!”

As this is a science blog, you may be surprised to learn that I am a German major. That I have been able to explore all of my different academic passions at Haverford is one reason why I have found my liberal arts education so rewarding. I learned German at Haverford, while taking courses like Chem225 and Bio200, which deepened my love for scientific inquiry as a means to better understanding ourselves and the world around us.

I have taken my knowledge of these different disciplines with me to Penn’s Center for Cognitive Neuroscience, where I am spending my summer in the lab of Anjan Chatterjee, MD (Class of 1980). Under the direct guidance of Anja Jamrozik, PhD, I am studying the neural bases of preposition metaphors. I am taking an interdisciplinary approach to my project here (which is super cool!).

I hope you will join me as I embark on this new adventure! But before we get started, I will address a question you probably have—

WHY GERMAN?
As a child, I spent countless hours bonding with my dad over soccer. In 2006, the German national team first caught my eye. I knew that they were something special, and it was only a matter of when they would become world champions (and eight years later, they finally did!). The more interviews I watched of my favorite players, the more curious I became about that beautiful language they were speaking.

So I made learning German one of my goals at Haverford, which proved a real challenge at times. I had to wrap my head around the fact that German nouns have genders, but at least I could fall back on memorizing them. Prepositions were different. Their meanings changed depending on context. For the notorious two-way prepositions like “auf,” the presence of motion in a sentence dictated what the article and adjective ending preceding the prepositional object were. Yet it was these very complexities of the German language that also fascinated me. I had to rethink my way of experiencing the world, and I loved it. I therefore decided to declare German as my major.

Before I move onto my discussion of my research project, however, I should mention that learning the German language is only one aspect of German studies. In the past three years, my love for German culture has grown immensely; and I have Professors Ulrich Schönherr, Imke Brust, Brook Henkel, and Kathleen Wright, as well as the Gotthardt family in Berlin, to thank for that.

WHY RESEARCH ON PREPOSITION METAPHORS AND THE BRAIN?
When the topic of studying abroad came up in Intermediate German, Professor Schönherr reminded us that such an experience allows students to not only learn about another culture but also reflect on their own. I soon asked myself if I could apply his same comment to language. After all, as speaking and writing in German became more natural to me, my native language of English began to sound and look very odd. Why was it that in Professor Ashly Bennett’s class on the novel, we used phrases like “marry off” or “break up” to describe the characters’ relationships? Why was it that during finals week, when I asked my friends how everything was “coming along,” a common answer was that they were “pulling through?”

Never before had it occurred to me to think about how my professors, friends, and I used language to communicate with one another. We used preposition metaphors like “coming along” all the time. We immediately knew to interpret them abstractly rather than take them literally.

That all said, I am absolutely thrilled to be at the Center for Cognitive Neuroscience this summer. By studying preposition metaphors on a neural level, I hope to demonstrate that they are not just adornments to our language, and that such abstract language is more than just “useful” to us. It may actually be fundamental to how we communicate with one another as well as how we understand the world to be.

WHAT DOES “THE NEURAL BASES OF PREPOSITION METAPHORS” EVEN MEAN?
Now that you know more about my background and approach to this project, let’s talk science! I am investigating whether or not the brain uses the same areas to process spatial uses of prepositions (e.g. the cat is in the box) and abstract, metaphorical uses of prepositions (e.g. he is in love). There are also temporal uses of prepositions (e.g. it happened in one second) that are important to keep in mind.

I will be working closely with patients who have parietal-temporal lesions on their brains. If there are any patients who can correctly produce only abstract, metaphorical uses of prepositions but not spatial uses of prepositions (and vice versa), then we can conclude that a dissociation in the brain exists between the neural bases of these two kinds of prepositions.

In my first several weeks here, I read a number of studies and other scholarly works on the different ways in which we employ commonly used prepositions like “over” and “under.” I started with “Network analyses of prepositional meaning: Mirroring whose mind— the linguist’s or the language user’s?” by Sandra and Rice (1995) , and I recently finished reading “Well-Hidden Regularities: Abstract Uses of in and on Retain an Aspect of Their Spatial Meaning” by Jamrozik and Gentner (2015). This background information will help Dr. Jamrozik and me as we prepare tests for patients that will gauge their ability to use either kind of preposition. The findings from our tests will be applicable to our understanding of the uses of prepositions in the real world. For example, Coventry and colleagues have shown that we tend to use “above” and “below” when talking about where things are located, whereas we tend to use “over” and “under” when talking about how things relate to each other and fulfill the purposes we attribute to them (e.g. “the Viking is under/below the shield”; 2001). It therefore will be important for us to take into account such distinctions between prepositions when we prepare our tests.

Figure 10 from Coventry and colleagues' "The Interplay between Geometry and Function in the Comprehension of Over, Under, Above, and Below" (Journal of Memory and Language, 2001).

Figure 10 from “The Interplay between Geometry and Function in the Comprehension of Over, Under, Above, and Below” (Coventry et al., 2001).

My summer work so far has been quite a challenge, as I do feel I am learning an entirely new language. It is filled with terms and concepts like Coventry and Garrod’s (2005) “functional geometric framework” I never would have imagined could exist! There is still a lot of information for me to digest— and I cannot wait to be at the level where I can ask Dr. Jamrozik the kinds of questions that will really probe my understanding of the material before me.

That said, learning about the evolution of the different theories on our uses of prepositions certainly has been an adventure. This intersection between cognitive psychology and linguistics is unchartered territory for me. I have had a lot of fun building the foundational knowledge I need to really tackle this research question head-on.

One last thing before I leave you with an Auf Wiedersehen! for the next several weeks—

The story of the Class of 1970 is truly inspiring, as I learned on Alumni Weekend. I also learned from them that you should not have any fear in doing what you love. Loving what you do is doing what is right by you. Go where you think you are meant to be in life; your Haverford education will help you to get there.

Haverford College Class of 1970

 

Welcome to the INSC Summer Scholars blog!

We come from different class years and have a wide range of academic interests, but we are all aspiring scientists and hope to share with you what our summer research experiences have been like!

We are Claire Cote ‘17 (Biology major, Health Studies minor), Claire Dinh ’16 (German major), Amanda Fleming ’16 (Biology major, Health Studies minor), Kerilyn Godbe ’16 (Chemistry major, Biochemistry concentration), Cora Hersh ’16 (Physics major), and Arjun Khandelwal ’17 (Physics and Astronomy double major, Economics minor).

Our research topics are the following:
Claire C.: Effects of soy isoflavone chemoprevention and smoking abstinence on candidate gene methylation of head and neck squamous cell carcinoma (School of Public Health at the University of Michigan, Ann Arbor)
Claire D.: Neural bases of preposition metaphors (Center for Cognitive Neuroscience at the University of Pennsylvania)
Amanda: Investigating the effects of transcription factor RP58 on development of GABAergic neurons in the mouse cerebellum (Department of Neurosurgery at the University of Pennsylvania)
Keri: The effects of varying the length of the SgI peptide on its hydrogelation properties (Department of Chemistry, Haverford College)
Cora: Dark matter searches with NuSTAR high-energy x-ray telescope (Department of Physics, Haverford College)
Arjun: Detection efficiency of long-period exoplanets exhibiting few planetary transit signals and scaling of the efficiency with the amount of data (Department of Physics and Astronomy, Haverford College)

Happy reading!

(For more information about student research funding opportunities by the Integrated Natural Sciences Center, click here).