Designing proteins that can sequence DNA

Filed Under: BiotechMolE PhDNews

Q&A with Molecular Engineering Ph.D. student Sinduja Marx

May 14, 2020

Sinduja Marx is a 5th year molecular engineering Ph.D. student co-advised by David Baker, a professor of biochemistry and director of the Institute for Protein Design, and Jens Gundlach, a professor of physics. We recently spoke with Marx about her research and her experience in the Molecular Engineering (MolE) Ph.D. program. 

MolE Ph.D. student Sinduja Marx purifies proteins in the lab as part of her work to improve next-generation DNA sequencing. Dennis Wise / University of Washington

What brought you to the molecular engineering program at UW?

As an undergraduate, I was interested in both bioengineering as well as physics. While initially those two fields might seem very different, there is actually a whole field biophysics dedicated to applying the theories and methods of physics to understand how biological systems work. Biology is complex, but when you break it down to the molecular level, it becomes easier to study. For example, fundamental electrostatic and hydrophobic interactions drive the structure and behavior of large molecules like DNA and proteins.

I knew that for grad school, I wanted to apply molecular-level science to the design of new devices, processes, or technologies. At the time, there were only two programs in the country dedicated to molecular engineering. I liked that the MolE program's core curriculum provided a solid foundation for working with molecules as well as the flexibility to choose additional courses that matched my own research interests. I also really appreciated the entrepreneurial environment at the UW and the many different opportunities and programs for innovators on campus.

You worked in industry before coming to UW. What was that experience like? How did it help prepare you for graduate school?

I had originally intended to go to grad school right after undergrad, but I ended up taking a two-year detour to work at Illumina, a widely known DNA sequencing company headquartered in San Diego. As a member of a small research and development team at Illumina, I worked with biochemists, physicists, mechanical engineers and software programmers to put together a next-generation sequencing device. My research there primed me for some of the questions I wanted to explore in my graduate work.

Tell us about your research.

Nanopore protein structures with three different sized pores
De novo design for nanopores of custom size. Image courtesy of Sinduja Marx

I am working on advancing the use of nanopore DNA sequencing technology. Unlike conventional sequencing technologies, nanopore sequencers can read long stretches of DNA in real-time, which is really advantageous when it comes to assembling lengthy genomes of organisms that do not have a reference sequence. Compared to today's widely used sequencers, nanopore sequencing machines are tiny, require few reagents and can run on very little power, all of which means they have the potential to be the first truly portable DNA sequencers.

What are nanopores?

Nanopores are naturally occurring proteins that form nanometer sized holes in cell membranes, allowing ions or molecules to be transported into and out of cells. A single strand of DNA can pass through a nanopore, one nucleotide at a time. Each nucleotide blocks the current moving through the pore in a characteristic way and so, by measuring the changes in current we can figure out the sequence of nucleotides in the DNA strand.

The downside of nanopore sequencing is that it is not as accurate as traditional sequencing methods. The physical geometry of the nanopore can greatly affect the signal to noise ratio and therefore the sequencing error rate. Researchers in the field are taking a variety of approaches to try to solve this signal to noise problem. My approach has been to ask, what if we could figure out the exact pore geometry necessary to accurately sequence DNA and then design that protein?

Why make a nanopore from scratch? Why not modify an existing nanopore?

If you try to change an existing protein, you don't really know what each amino acid does and so, any changes you make may alter the protein's structure and affect its function. In contrast, when you design a protein from scratch, you know the role of every amino acid relative to the intended function. Engineering a protein from scratch however, is easier said than done. Engineering membrane proteins is particularly challenging because predicting a large protein's structure based on the amino acid sequence relies on having known structures of many similar proteins. Membrane proteins are difficult to crystallize, which means there aren't a lot of available structures in the protein database.

Starting from a first principles design model, my goal has been to simulate amino acid sequences that are predicted to fold into the desired channel shape, express them in bacterial cells, and then suspend them in artificial lipid bilayers to study their conductance properties and potential for DNA sequencing.

What has been the best part of grad school? What has been the most challenging?

The best part of grad school has definitely been the freedom to deeply explore and develop new ideas. For me, the most difficult part has been the high failure rate. This is of course expected in research, but it can be really taxing day in and day out. Along the way, I've learned a lot about how to convince people of my interpretations or decisions and how to receive criticism from advisors, committee members, etc.; both useful skills to have wherever my career takes me next.

What advice do you have for future students?

There are plenty of important problems that don't make great Ph.D. projects. Grad students must quickly learn to evaluate whether they have a reasonable line of attack for the problem, and if not, be ready to move on to the next important problem. I highly recommend reading Richard Hamming's talk "You and your research" where he expands on this point.

What do you want to do next?

Honestly, right now I'm not sure. I've always seen myself as a research scientist, but I would like to spend some time exploring all of the potential career paths available to me. I know there are a lot of other ways outside of the lab that I can use my scientific expertise, but at this point I still need more information to figure out which path is the right one for me.

To learn more about the MolE Ph.D. program visit