About

Philip Bevilacqua is a Distinguished Professor of Chemistry and Biochemistry and Molecular Biology at Penn State University, where he also serves as Co-Director of the Center for RNA Molecular Biology. His research focuses on RNA folding in vivo and genome-wide, RNA regulation of gene expression, ribozyme mechanisms, and the roles RNA may have played in the emergence of life on early Earth. Bevilacqua's work involves developing methods to identify and characterize modifications to RNA structure that can alter its function and serve as potential drug targets. His contributions include advancing understanding of RNA structure probing, RNA thermometers that regulate translation, and the molecular principles underlying RNA's versatility and functionality.

Research topics

• Biology
• Chemistry
• Biochemistry
• Biophysics
• Cell biology
• Genetics
• Organic chemistry
• Molecular biology
• Chromatography
• Ecology
• Chemical engineering
• Thermodynamics

Selected publications

• Optimized tRNA structure–seq reveals robust tRNA secondary structures in <i>S. cerevisiae</i> under mild stress conditions

  bioRxiv (Cold Spring Harbor Laboratory) · 2026-02-25

  articleOpen access

  Abstract RNA structure plays a crucial role in diverse biological processes beyond the translation of genetic information. Therefore, the development of reliable methods for RNA structure prediction is essential for understanding RNA structure–related functions, however accurate and comprehensive RNA structure prediction remains challenging. Here, we focus on secondary structure prediction of transfer RNA (tRNA) using structure probing coupled with next–generation sequencing (tRNA Structure–seq). In silico prediction of Saccharomyces cerevisiae tRNA secondary structures achieves only 56.9% accuracy on average. Incorporation of dimethyl sulfate (DMS) probing data improve prediction accuracy to 87.4%, which is still not sufficient for practical tRNA structure prediction. To overcome this, we optimized the tRNA Structure–seq analysis pipeline by explicitly incorporating natural tRNA modifications detected in tRNA sequencing data and by refining pseudo–free energy parameters specifically optimized for tRNA structure prediction. Using this optimized pipeline, the average prediction accuracy is remarkably improved to 94%. Furthermore, analysis of multiple structural conformations predicted from DMS probing data indicates that S. cerevisiae tRNAs predominantly adopt the canonical cloverleaf secondary structure under in vivo conditions. Finally, we examined tRNA structures under mild stress conditions, including heat stress, osmotic stress, and antibiotic stress. These perturbations had minimal effects on in vivo tRNA secondary structure, demonstrating that S. cerevisiae tRNAs maintain structural stability under physiologically relevant stress conditions. In summary, our results establish an optimized tRNA Structure–seq analysis that enables highly accurate tRNA secondary structure prediction and reveals the intrinsic robustness of tRNA structures in living cells.

  PublisherDOI

• RNA–peptide–DNA together from the beginning

  Nature Chemical Biology · 2026-02-18

  articleSenior author
  PublisherDOI

• Nearest Neighbor Parameters for Estimating the Folding Stability of RNA Including Pseudouridine

  bioRxiv (Cold Spring Harbor Laboratory) · 2026-05-17

  articleOpen access

  Nearest neighbor parameters are widely used in software for estimating the conformational stability of an RNA sequence folding into a specific structure. Folding stability for RNA with canonical nucleotides A, C, G, and U has been widely studied, but the same is not true for most modified nucleotides. In this work, we present a comprehensive set of nearest neighbor parameters for estimating the folding stability of RNA including pseudouridine in helical or loop contexts. These parameters are derived from 210 optical melting experiments involving helices with pseudouridine-A and pseudouridine-G pairs and with pseudouridine in loop motifs. The experiments include sequences with pseudouridine and U in the same strand, including U-A and U-G pairs, allowing us to consider the folding stability of sequences with both U and pseudouridine. On average, pseudouridine stabilizes RNA folding compared to U in an analogous motif, although this effect is sequence-context dependent. These parameters improve the modeling of folding stability for RNA secondary structures containing pseudouridine. We demonstrate that these parameters successfully model the secondary structure change for Saccharomyces cerevisiae U2 snRNA when two additional inducible pseudouridines are present. These parameters are freely available and incorporated into the RNAstructure software package.

  PublisherDOI

• RNA Folding Nearest Neighbor Parameters Including the Modification 1-Methyl-Pseudouridine

  bioRxiv (Cold Spring Harbor Laboratory) · 2026-04-11

  articleOpen access

  Nearest neighbor analysis is commonly used to estimate RNA folding stabilities. In this contribution, we report a set of RNA folding nearest neighbor parameters for estimating free energy change for RNA sequences including 1-methyl-pseudouridine. Development of mRNA vaccines has identified 1-methyl-pseudouridine as a key nucleobase modification for suppressing innate immune responses. However, the contributions of these modifications to RNA folding stability were unclear. Our new parameters provide helical terms for 1-methyl-pseudouridine-adenine and 1-methyl-pseudouridine-guanine base pairs. The parameters also estimate loop stabilities for loops with 1-methyl-pseudouridine or a combination of 1-methyl-pseudouridine and uridine. These parameters are derived using 208 optical melting experiments and tested against an additional 16 optical melting experiments. On average, we find that substitution of uridine with 1-methyl-pseudouridine stabilizes RNA folding, with the extent of stabilization depending on adjacent sequence. The estimation of tRNA folding ensembles for tRNA sequences with 1-methyl-pseudouridine was significantly improved using the new nearest neighbor parameters. The new nearest neighbor parameters are provided as part of the RNAstructure software package. With these parameters, the secondary structures of natural sequences with 1-methyl-pseudouridine and mRNA therapeutics fully substituted with 1-methyl-pseudouridine can be modeled.

  PublisherOA PDFDOI

• VariantFoldRNA: a flexible, containerized, and scalable pipeline for genome-wide riboSNitch prediction

  NAR Genomics and Bioinformatics · 2025-03-29

  articleOpen access

  Single nucleotide polymorphisms (SNPs) can alter RNA structure by changing the proportions of existing conformations or leading to new conformations in the structural ensemble. Such structure-changing SNPs, or riboSNitches, have been associated with diseases in humans and climate adaptation in plants. While several computational tools are available for predicting whether an SNP is a riboSNitch, these tools were generally developed to analyze individual RNAs and are not optimized for genome-wide analyses. To fill this gap, we developed VariantFoldRNA, a flexible, containerized, and automated pipeline for genome-wide prediction of riboSNitches. Our pipeline automatically installs all dependencies, can be run locally or on high-performance clusters, and is modular, enabling the user to customize the analysis for the research question of interest. VariantFoldRNA can predict riboSNitches genome-wide at user-specified temperatures and splicing conditions, opening the door to novel analyses. The pipeline is an open-source command-line tool that is freely available at https://github.com/The-Bevilacqua-Lab/variantfoldrna.

  PublisherDOI

• BPS2025 - Phase-separated aqueous polyelectrolyte solutions as “proto-cytoplasm” and their response to guest molecules and environmental perturbations

  Biophysical Journal · 2025-02-01

  article
  PublisherDOI

• Identification and characterization of shifted G•U wobble pairs resulting from alternative protonation of RNA

  bioRxiv (Cold Spring Harbor Laboratory) · 2025-01-02 · 2 citations

  preprintOpen accessSenior authorCorresponding

  Abstract RNA can serve as an enzyme, small molecule sensor, and vaccine, and it may have been a conduit for the origin of life. Despite these profound functions, RNA is thought to have quite limited molecular diversity. A pressing question, therefore, is whether RNA can adopt novel molecular states that enhance its function. Covalent modifications of RNA have been demonstrated to augment biological function, but much less is known about non-covalent alterations such as novel protonated or tautomeric forms. Conventionally, a G•U wobble has the U shifted into the major groove. We used a cheminformatic approach to identify four structural families of shifted G•U wobbles in which the G instead resides in the major groove of RNA, which requires alternative tautomeric states of either base, or an anionic state of the U. We provide experimental support for these shifted G•U wobbles via the potent, and unconventional, in vivo reactivity of the U with dimethylsulfate (DMS) in three organisms. These shifted wobbles may play important functional roles and could serve as drug targets. Our cheminformatics approach is general and can be applied to identify alternative protonation states in other RNA motifs, as well as in DNA and proteins. Graphical Abstract

  PublisherOA PDFDOI

• Prevalence of dual-donating amines in key regions of functional RNAs

  bioRxiv (Cold Spring Harbor Laboratory) · 2025-06-05

  preprintOpen accessSenior authorCorresponding

  RNA performs many critical functions nearly all of which are enabled by complex hydrogen bonded structures. Nucleotides possess far fewer hydrogen bond donors than acceptors, and only the exocyclic amine can donate two H-bonds, suggesting a specialized role. To assess the prevalence and structural contexts of dual-donating amines within structured RNAs, we created a computational workflow that mines and analyzes experimental RNA-containing structures. We evaluated H-bonding in over 250,000 amines from more than 1,800 structures. Dual-donating amines were found most frequently in G's where they regularly interacted with diverse pairs of acceptors. In contrast, the dual-donating amines of A's and C's were less frequent and they interacted with a more select set of acceptors. For all three nucleobases, amines that were dual- donating had both reduced solvent accessibility and higher atom density relative to amines that were non-donating, indicating a tendency of dual donors to be more buried and help compact the RNA. Moreover, analysis of RNA pseudo-torsion angles revealed that dual-donating amines are enriched in two non A-form conformations, both of which are present in S-motifs found in the sarcin-ricin loop of rRNA. We find that dual-donating amines populate additional structural motifs including the GNRA tetraloop receptor, the kink-turn, and the WC/H A-minor motif, which are present in the self-splicing group I intron, the SAM riboswitch, and the poly(A)-bound ENE. We suggest that dual-donating amines may enhance interactions by reducing conformational entropy loss as well as strengthening nearby H-bonds.

  PublisherOA PDFDOI

• Cryo-EM study and in vivo chemical mapping of the Methanosarcina acetivorans ribosome and its dimerization via a repurposed enzyme and translation factor

  Journal of Biological Chemistry · 2025-09-04 · 2 citations

  articleOpen access

  Despite the overall conservation of ribosomes across all domains of life, differences in their 3D architecture, rRNA sequences, ribosomal protein composition, and translation factor requirements reflect lineage-specific adaptations to environmental niches. In the domain Archaea, structural studies have primarily focused on nonmethanogenic thermophiles and halophiles, leaving it unclear whether these represent the broader Archaea domain. Here, we report the cryo-electron microscopy (cryo-EM) structure of the ribosome from Methanosarcina acetivorans, a previously unreported high-resolution structure from a model mesophilic methanogenic archaeon. Compared to ribosomes from extremophiles, the M. acetivorans ribosome has a simplified architecture, lacking paralogous duplications and containing a reduced complement of ribosomal proteins. Structures of the large subunit (50S) from cells grown with either methanol or acetate show conserved rRNA folding and protein composition. High-resolution structures of the 50S subunit from the two growth substrates enabled us to investigate structural properties that may influence in vivo dimethyl sulfate reactivity, an orthogonal chemical approach used to probe RNA structure. We observed good agreement between in vivo dimethyl sulfate reactivity and ribosome structure. Finally, we identify a previously uncharacterized ribosome dimerization mode involving only 50S subunits and mediated by a heterotetrameric complex of PurH and aEF2-proteins with alternative metabolic and translational roles. This macromolecular assembly, which we term the methanogen ribosome dimerization factor, likely mediates ribosome hibernation, revealing an alternative regulatory mechanism in translation.

  PublisherOA PDFDOI

• Identification and characterization of shifted G•U wobble pairs resulting from alternative protonation of RNA

  Nucleic Acids Research · 2025-06-07 · 3 citations

  articleOpen accessSenior author

  RNA can serve as an enzyme, small molecule sensor, and vaccine, and it may have been a conduit for the origin of life. Despite these profound functions, RNA is thought to have limited molecular diversity. A pressing question is whether RNA can adopt novel molecular states that enhance its function. Covalent modifications of RNA have been demonstrated to augment biological function, but much less is known about non-covalent alterations such as novel protonated or tautomeric forms. Conventionally, a G•U wobble has the U located in the major groove. We used a cheminformatic approach to identify four structural families of shifted G•U wobbles in which the G instead resides in the major groove, which requires alternative tautomeric states of either base, or an anionic state of the U. We provide experimental support for these shifted G•U wobbles via the unconventional in vivo reactivity of the U with dimethylsulfate (DMS). These shifted wobbles may play functional roles and could serve as drug targets, as they are common in Bacteria and chloroplasts, but underrepresented in Eukaryotes and Archaea. Our cheminformatics approach can be applied to identify alternative protonation states in other RNA motifs, as well as in DNA and proteins.

  PublisherOA PDFDOI

Recent grants

• Collaborative Research: RESEARCH-PGR: Genetic and environmentally-induced functional variation in the rice RNA structurome

  NSF · $1.8M · 2021–2026

• Mechanistic studies of proton transfer in ribozyme self-cleavage

  NSF · $356k · 2012–2017

• NIH Grant R01GM058709

  NIH · $3.0M · 2013

• The Influence of Secondary Structure on the Folding and Catalysis of Functional RNAs

  NSF · $908k · 2005–2010

• MRI: Acquisition of High-Throughput Calorimeters for Ligand-Biopolymer Discovery and Characterization

  NSF · $460k · 2009–2012

Frequent coauthors

• Sarah M. Assmann

  Pennsylvania State University

  42 shared

• David J. Proctor

  41 shared

• Ryszard Kierzek

  Institute of Bioorganic Chemistry, Polish Academy of Sciences

  30 shared

• Joshua M. Blose

  26 shared

• Barbara L. Golden

  Purdue University System

  23 shared

• Elżbieta Kierzek

  23 shared

• Durga M. Chadalavada

  Pennsylvania State University

  21 shared

• Narayanan Veeraraghavan

  Rady Children's Hospital-San Diego

  19 shared

Labs

• Bevilacqua LabPI

Awards & honors

• Early Career Award from Society of Biological Inorganic Chem…
• Gordon Hammes Scholar Award (2019)