Kylie J. Walters, Ph.D.

Kylie J. Walters, Ph.D.
Senior Investigator
Head, Protein Processing Section

Dr. Walters is an internationally renowned structural biologist who has made seminal discoveries of how targeted protein degradation occurs in cells. Her laboratory has applied a multidisciplinary approach that includes cutting-edge cellular and structural biology methods, including CRISPR-based gene editing, NMR spectroscopy, and cryoelectron microscopy, to uncover how the proteasome recognizes and processes its substates. Major discoveries include identifying Rpn1 and Rpn13 as proteasome substrate receptors as well as a binding site for E3 ubiquitin ligase E6AP/UBE3A at the proteasome in Rpn10. These findings have led to new therapeutic strategies for cancer which she is pursuing by using structure-based drug design.

Areas of Expertise

1) structural biology, 2) structure-based drug design, 3) ubiquitin signaling, 4) proteasome, 5) NMR spectroscopy, 6) cryoelectron microscopy

Contact Info

Kylie J. Walters, Ph.D.
Center for Cancer Research
National Cancer Institute
Bldg. 538/Room 167
Frederick, MD 21702
Ph: 301-228-4374

The Walters lab studies the structural and mechanistic basis of ubiquitin signaling events, proteasome function, and protein quality control. We use an interdisciplinary approach that merges functional cell-based assays with a variety of biophysical and structural biology techniques to define the mechanistic underpinnings of how protein lifetimes are regulated in cells. We have helped to identify the mechanisms of substrate recognition by the proteasome, identifying Rpn1 and Rpn13 as substrate receptors that bind to ubiquitin chains directly and to the shuttle factors that deliver ubiquitinated substrates to the proteasome. By using NMR spectroscopy, we have solved structures of these receptors complexed with ubiquitin chains and shuttle factors. We have also used cryoelectron microscopy to study ubiquitin chain binding to the 26S proteasome.

We discovered that Rpn13 is targeted in its ubiquitin-binding domain by cell permeable molecules that restrict cancer cell proliferation. We generated CRISPR cell lines that have revealed the importance of Rpn13 at the proteasome and its requirement for triggered apoptosis by these small molecules. Small molecules that inhibit the catalytic core particle of the proteasome are approved for treatment of hematological cancers and the targeting of Rpn13 is expected to function synergistically with these core particle inhibitors. We are further developing other Rpn13-targeting molecules by using structure-based drug design methods.

In a related project, we recently identified a binding site for the E3 ubiquitin ligase E6AP/UBE3A at the proteasome in substrate receptor Rpn10. This functional domain is the first found at the proteasome to recruit ubiquitination machinery. E6AP is targeted by the human papilloma virus E6 oncoprotein leading to degradation of tumor suppressor p53 and its dysfunction is also associated with prostate cancer, Angelman syndrome, and autism spectrum disorders. We continue to study the role of E6AP at the proteasome as well as other protein-protein interactions in the ubiquitin-proteasome pathway to understand the determinants of protein targeting for degradation as well as the mechanistic features that go awry in these pathways during carcinogenesis and neurological disorders.

NIH Scientific Focus Areas:
Biomedical Engineering and Biophysics, Cancer Biology, Molecular Biology and Biochemistry, Molecular Pharmacology, Structural Biology
View Dr. Walters' PubMed Summary.

Selected Key Publications

  1. Shi Y, Chen X, Elsasser S, Stocks BB, Tian G, Lee BH, Shi Y, Zhang N, de Poot SA, Tuebing F, Sun S, Vannoy J, Tarasov SG, Engen JR, Finley D, Walters KJ
    Science. 351: pii: aad9421, 2016. [ Journal Article ]
  2. Buel GR, Chen X, Chari R, O'Neill MJ, Ebelle DL, Jenkins C, Sridharan V, Tarasov SG, Tarasova NI, Andresson T, Walters KJ
    Nat. Commun. 11: 1291, 2020. [ Journal Article ]
  3. Lu X, Ebelle DL, Matsuo H, Walters KJ
    Structure. 28:, 2020. [ Journal Article ]
  4. Lu X, Nowicka U, Sridharan V, Liu F, Randles L, Hymel D, Dyba M, Tarasov SG, Tarasova NI, Zhao XZ, Hamazaki J, Murata S, Burke TR Jr, Walters KJ
    Nat. Commun. 8: 15540, 2017. [ Journal Article ]
  5. Chen X, Dorris Z, Shi D, Huang RK, Khant H, Fox T, de Val N, Williams D, Zhang P, and Walters KJ
    Structure. 28:, 2020. [ Journal Article ]

Dr. Walters’ training in the field of biophysics began as an undergraduate at Wesleyan University, where she obtained HHMI funding to apply NMR methods to study the dynamic properties of DNA under the supervision of Dr. Irina Russu. She then obtained a Ph.D. in Biophysics from Harvard University, where with Dr. Gerhard Wagner, she developed and used methods to study protein structure, dynamics, and interaction mechanisms. As an American Cancer Society Postdoctoral Fellow with Dr. Peter Howley in the Pathology Department at Harvard Medical School, she began to apply structural biology methods to study the ubiquitin-proteasome pathway. In 2002, Dr. Walters joined the University of Minnesota as an Assistant Professor and was promoted to Associate Professor with tenure in 2008. During this time, Dr. Walters was selected for an American Cancer Society Research Scholar award for her work in protein quality control. In 2013, she moved to the Center for Cancer Research as a Senior Investigator, where she runs a vibrant research program that aims to apply basic science discoveries to translational benefit in the clinic. She has been awarded an NIH Director’s Award, NIH OD Honor Award, NCI Champions-Leading Diversity Award, and CCR top scientific advance. She currently serves on the CCR Science Board, Editorial Board of PLOS Biology, Advisory Board for the journal Structure, and has served as Guest Editor for PNAS and Current Opinions in Structural Biology.

Name Position
Gwen Buel Ph.D. Postdoctoral Fellow (CRTA)
Xiang Chen, Ph.D. Staff Scientist
Xiuxiu Lu Ph.D. Research Fellow (Visiting)
Vasty Osei Amponsa Ph.D. Postdoctoral Fellow (Visiting)
Brandon J. Wright Ph.D. Postdoctoral Fellow (CRTA)
Anne Kaplan Postdoctoral Fellow 09/03/2019-10/31/2020

Current position: Research Scientist, Aurobindo

Olumide Kayode Postdoctoral Fellow 03/2017-09/2020

Current position: Revolve Biotechnologies

Anthony Cruz Postbaccalaureate Student 09/2019-09/2020

Current position: Undergraduate Researcher at Univeristy of Miami, FL

Zachary Dorris Werner H. Kirsten Student Intern 06/2019-05/2020

Current position: Undergraduate Student at University of Maryland, College Park

Danielle Ebelle Postbaccalaureate Student 06/2016-06/2019

Current position: Medical Student at Saba University School of Medicine

Vinidhra Sridharan Postdoctoral Fellow 8/2016-11/2018

Current position: Scientist at Bristol-Myers Squibb


Evan Hooper Student 6/2017-5/2018

Current position: Undergraduate Student at Florida State University


Fahu He Research Fellow (VP) 6/2013-11/2016

Current position: NMR Facility Manager, St. Louis University


Urszula Nowicka Postdoctoral Fellow 7/2014-11/2016

Current position: Scientific Advisor, Center of New Technologies, University of Warsaw


Sarah Durham Student 6/2015-8/2016

Current position: Undergraduate student at Lebanon Valley College


Fen Liu, Ph.D. Research Fellow (VP) 6/2013-7/2016

Current Position: Principle Scientific Consultant, 100BioTech, Co. Ltd.


Leah A. Randles, Ph.D. Postdoctoral Fellow; Graduate Student 01/2013-05/2016; 2/2007-12/2012

Current position: Chemist, Syntiron


Jacob Vannoy Student 6/2014-8/2015; 3/2017-8/2017

Current position: Student at University of Virginia


Morgan Hoffman Student 06/2014-08/2015

Current position: Undergraduate student at University of Maryland, College Park


Yanhong Shi, Ph.D. Sabbatical Visiting Professor 7/2013-8/2014

Current position: Faculty at Shanghai Institute of Materia Medica


Aaron C. Ehlinger, Ph.D. Graduate Student 2/2008-7/2013

Current position: Postdoctoral Fellow at Vanderbilt University


Molly Westfield High School Researcher summer 2012

Current position: Undergraduate student at the University of Minnesota


Kimberly Wadsworth, Ph.D. Postdoctoral Fellow 5/2009-5/2011

Current position: Senior Immunogenetic Specialist at Be The Match operated by National Marrow Donor Program


Yang Kang, Ph.D., DDS PhD Grad Student and Postdoctoral Fellow 9/2003-6/2010

Current position: Assistant Clinical Professor at University of New England, Maine


Naixia Zhang, Ph.D. Research Assistant Professor 5/2004-2/2010

Current position: Faculty at Shanghai Institute of Materia Medica


William Casey Solomon, Ph.D. Undergraduate Student Researcher 1/2008-6/2009

Current position: Visiting Assistant Chemistry Professor, St Olaf College


Cheryl Ethan, Ph.D. Research Specialist 3/2007-6/2007

Current position: Research Scientist, R&D Systems Inc.


Qinghua Wang, Ph.D. PhD Grad Student and Postdoctoral Associate 1/2002-9/2006

Current position: Instructor, University of Delaware


Nathan Winter, Ph.D. Sabbatical Visiting Professor 1/2004-6/2004

Current position: Professor at St Cloud State University Department of Chemistry


Nov. 2020: Congratulations to Dr. Xiang Chen for his publication of how unanchored ubiquitin chains bind the proteasome – so-entering the world of cryoEM!

Congratulations to Dr. Anne Kaplan for getting a position at Aurobindo; we will miss you!

Congratulations to Dr. Olumide Kayode for getting a position with Revolve Biotechnologies; you will be missed!

August 2020: Congratulations to lead author Vasty Osei-Amponsa for making the cover of MCB for her studies of Rpn13 Pru domain and UCHL5 requirement in the clearance of ubiquitinated proteins and RA190 targeting.

May 5th 2020: Congratulations to Xiuxiu Lu and Danielle Ebelle on revealing the mechanism of how hRpn13 binds K48 ubiquitin chains, published today in Structure, with an excellent commentary by Jeroen Roelofs.

March 10th 2020: Congratulations to Gwen Buel and Xiang Chen on their discovery of a binding site for E6AP at the proteasome, published today in Nature Communications.

Members of the group celebrate achievements of 2019 at a restaurant in downtown Frederick.

Members of the group celebrate achievements of 2019 at a restaurant in downtown Frederick.

Congratulations to Dr. Xiuxiu Lu for receiving a Best Poster Award at the 2018 NCI Structural Biology Retreat

Structural Biophysics Laboratory 2018 Retreat - Poster session

Congratulations to Dr. Xiang Chen and Dr. Kylie Walters for receiving CCR Federal Technology Transfer Awards

Congratulations to Dr. Gwen Buel for being selected as a winner of NIH Fellows Award for Research Excellence (FARE) 2019 competition.

Congratulations to Dr. Xiang Chen for being selected as a winner of NIH Fellows Award for Research Excellence (FARE) 2017 competition. Xiang used this award to attend and present a poster at Cold Spring Harbor Laboratory Conference “The ubiquitin Family” on April 18-22, 2017.

Congratulations to Dr. Xiuxiu Lu for being selected to give an oral presentation at the ASBMB 2017 meeting in Chicago on April 22-26.

Dr. Vinidhra Sridharan presented a poster at the Cold Spring Harbor Laboratory Conference “The Ubiquitin Family”, April 18-22, 2017.

The Protein Processing Section pose for a photo during the summer of 2018

The Protein Processing Section pose for a photo during the summer of 2018.