Never Stop Exploring: Spotlight on Squire Booker
As a young scientist, Squire Booker, Ph.D., wanted to explore anything and everything—from distant worlds in space to microscopic worlds on Earth. Now, as the Evan Pugh University professor of chemistry and of biochemistry and molecular biology and the Eberly Family Distinguished Chair in Science at The Pennsylvania State University, Booker wants his students to have the same opportunity to explore their passions in STEM, and, ultimately, pursue projects that they are curious and excited about.
Booker grew up in east Texas on the border of Texas and Louisiana. His uncle had previously worked for the National Aeronautical and Space Administration (NASA) and would often take Booker to NASA in nearby Houston to explore the exhibits. “When I was a kid, the Apollo mission was going on, so we would watch the launches and witness the astronauts coming back from moon missions,” Booker recalled. “My first interest in science was astronomy.”
At the same time, another uncle, who taught mathematics, inspired Booker’s curiosity for solving complex problems and puzzles. “I drifted toward subjects that combined math with science,” Booker said. As a result, he selected chemistry, a subject that combined these 2 interests, as his major at Austin College in Sherman, Texas. In 1986, during his time as an undergraduate student, Booker was part of the first cohort for Massachusetts Institution of Technology (MIT)’s Minority Summer Science Research Program. During this program, Booker’s research focused on the purification of methyl viologen (MV) hydrogenase, an enzyme isolated from Methanobacterium thermoautotrophicum. This was also when he first became interested in the intersection between chemistry and microbiology.
The role of MV hydrogenase was unknown at the time; however, mechanistically, it could remove the electrons from hydrogen gas and pass them on to methyl viologen, a type of organic compound that is sometimes used as an herbicide. Methyl viologen turns a deep violet color upon reduction, which provides an easy method to observe the reaction. Through his experience working with MV hydrogenase, Booker became familiar with protein purification and anaerobic setups. He also developed an appreciation for the research process—from designing an experiment to working toward a specific goal.
Following his involvement with MIT’s summer research program, Booker completed his undergraduate studies and elected to pursue a Ph.D. in biochemistry at MIT. That was in 1987. Six years later, in 1993, Booker co-authored the paper, “Cloning, sequencing, and expression of the adenosylcobalamin-dependent ribonucleotide reductase from Lactobacillus leichmannii,” with his Ph.D. advisor, JoAnne Stubbe, Ph.D.
Ribonucleotide reductases are enzymes that are integral to DNA replication, as they catalyze the rate limiting step in DNA biosynthesis. This study looked at the first sequence of a ribonucleotide reductase that did not use an iron-containing cofactor (i.e., a non-protein chemical compound that helps an enzyme act as a catalyst). Instead, the ribonucleotide reductase of L. leichmannii depends on adenosylcobalamin. “This led to some very important studies on how adenosylcobalamin functions in enzyme catalysis, in general, and in ribonucleotide reductases, in particular,” Booker added.
In 1994, Booker became a NSF-NATO Postdoctoral Fellow and investigated inducible nitric oxide synthase at Université de Paris V (René Descartes). Nitric oxide, which was named molecule of the year in 1992, is a gaseous molecule involved in signaling pathways that lead to vasodilation. “Nitric oxide is generated from arginine, and everyone in the world, at the time, was trying to figure out how [it’s generated],” Booker said. When Booker returned to the U.S. the following year, he embarked on a second postdoc at the Enzyme Institute, University of Wisconsin-Madison, where he worked alongside advisor Perry Frey, Ph.D.
As an NIH Postdoctoral Fellow, Booker probed at iron-sulfur centers in lysine 2,3-aminomutase, eventually co-authoring the paper, “S-Adenosylmethionine-dependent reduction of lysine 2,3-aminomutase and observation of the catalytically functional iron-sulfur centers by electron paramagnetic resonance,” in 1998. Radicals are chemicals that have unpaired electrons and are useful in chemical studies because they can serve as intermediates. At the time, there was a new radical-generating system that mimicked adenosylcobalamin (also known as coenzyme B12) that Booker had studied in Stubbe’s lab. But instead of using adenosylcobalamin, it generated the same kind of radical using S-adenosylmethionine. These enzymes now belong to a super family of enzymes called radical S-adenosylmethionine (SAM). “That’s what drew me to this area of research in [Dr. Frey’s] lab,” Booker added. “He was really the main founder of radical essence of adenosylmethionine molecules.”
While it was known that radical SAM enzymes require iron-sulfur clusters to function, it wasn’t clear what exactly the clusters were doing. Booker and his team showed that only when the cluster is in a reduced state can SAM be cleaved to generate the 5'-deoxyadenosyl 5'-radical. “This enzyme was isolated from Clostridium subterminale, an obligate anaerobe,” Booker said, explaining that SAM enzymes help anaerobic organisms conduct chemical reactions that would necessitate oxygen in aerobes. "There is an anaerobic counterpart to ribonucleotide reductase, which has a radical SAM component. It is believed that this anaerobic enzyme was the key to the evolution of life before oxygen arrived in the atmosphere.”
It was through this work, studying the enzyme lysine, 2,3-aminomutase (a type of radical SAM enzyme) in 1998, that Booker started to observe deeper connections between microbiology and chemistry. Booker discovered a number of interesting reactions in bacteria. He focused on understanding how certain bacteria, like Staphylococcus aureus acquire the cfr gene and put a methyl group on their ribosomal RNA.
“The enzymes that are capable of doing this reaction cause resistance to more than 5 classes of antibiotics,” Booker explained, emphasizing the public health implications of this type of research. “Most of our antibiotics actually target ribosomes, so these are 5 classes of clinically relevant antibiotics.” Developing compounds that inhibit this enzyme could help protect the current “arsenal of antibiotics.” Additionally, further study of the key steps in the biosynthesis of various antibiotics could assist in the development of more effective drugs.
Still, Booker reflected that investigating this hypothesis was not a straightforward journey. “Each time we did an experiment, we got an answer that we didn't predict. The mechanism sort of gradually evolved,” Booker said. “It's really fun when you get exciting and unexpected results.” Here, Booker was reminded that when you encounter failure, or unexpected results, it’s an opportunity to learn rather than a shortcoming. "You learn from failure. What you don't learn from is not doing anything,” Booker said. “If you do something, and you fail, then you learn—either it was not the right approach, or the hypothesis wasn’t right. You get information back.”
Currently, Booker continues to set his sights on RNA methylation and how microbes make certain natural products that serve as antibiotics. One of the steps that is often found in many antibiotic biosynthetic pathways is the methylation of inert carbon, (i.e., carbon that you cannot typically do acid-base chemistry with). “[My team] were among the first people to show that these radical SAM enzymes can actually perform radical dependent methylation,” he shared.
The ability to explore numerous avenues in STEM is now an experience Booker hopes to return to his students. In particular, Booker strives to provide early-career scientists with as many research opportunities as possible. In addition to working with graduate students and postdocs, Booker has trained more than 60 undergraduate students, many of whom have published research as his mentees. He also trained 2 high school students, one of whom is now a postdoc at Stanford University, and the other is currently finishing his graduate studies at MIT.
“I always look for passion,” he said, “One of my advisors once told me there are better jobs than being a scientist, because you typically worked long hours and the pay wasn’t always great—you did this because you were passionate about science, you really enjoyed figuring out problems and dealing with puzzles. I look for that in all the people I work with, and I try to find projects for them to engage with that are at the forefront of science so they can be excited about what they’re doing.”
Booker noted that compared to his time as an undergraduate student, today, there are many more organizations that aim to support students from historically underrepresented groups in STEM, which he encourages his own students to engage with as much as possible. Booker previously served on the Minority Affairs Committee for the American Society of Biochemistry and Molecular Biology (ASBMB). He is currently part of the steering committees for the American Biomedical Research Conference for Minoritized Students (ABRCMS) and ASBMB MOSAIC. “To future scientists from diverse backgrounds, I would say stay away from people who are negative, and drift toward people who are positive and who hold you up rather than put you down.”