Theresa A Ramelot

Rensselaer Polytechnic Institute | RPI · Center for Biotechnology and Interdisciplinary Studies

IscU is a highly conserved protein that serves as the scaffold for IscS-mediated assembly of iron-sulfur ([Fe-S]) clusters. We report the NMR solution structure of monomeric Haemophilus influenzae IscU with zinc bound at the [Fe-S] cluster assembly site. The compact core of the globular structure has an alpha-beta sandwich architecture with a three...

The structure of human protein HSPC034 has been determined by both solution nuclear magnetic resonance (NMR) spectroscopy and X-ray crystallography. Refinement of the NMR structure ensemble, using a Rosetta protocol in the absence of NMR restraints, resulted in significant improvements not only in structure quality, but also in molecular replacemen...

While structural symmetry is a prevailing feature of homo-oligomeric proteins, asymmetry provides unique mechanistic opportunities. We present the crystal structure of full-length TRAP1, the mitochondrial Hsp90 molecular chaperone, in a catalytically active closed state. The TRAP1 homodimer adopts a distinct, asymmetric conformation, where one prot...

Psilocybin, the prodrug of the psychoactive molecule psilocin, has demonstrated promising results in clinical trials for the treatment of addiction, depression, and post-traumatic stress disorder. The development of a psilocybin production platform in a highly engineerable microbe could lead to rapid advances towards the bioproduction of psilocybin...

There has been considerable recent progress in protein structure prediction using deep neural networks to predict inter-residue distances from amino acid sequences1,2,3. Here we investigate whether the information captured by such networks is sufficiently rich to generate new folded proteins with sequences unrelated to those of the naturally occurr...

Energetically favorable local interactions can overcome the entropic cost of chain ordering and cause otherwise flexible polymers to adopt regularly repeating backbone conformations. A prominent example is the α helix present in many protein structures, which is stabilized by i, i + 4 hydrogen bonds between backbone peptide units. With the increase...

RHOA mutations are found at diverse residues in various cancer types, implying mutation- and cell-specific mechanisms of tumorigenesis. Here, we focus on the underlying mechanisms of two gain-of-function RHOA mutations, A161P and A161V, identified in adult T-cell leukemia/lymphoma. We find that RHOAA161P and RHOAA161V are both fast-cycling mutants...

A general approach to design proteins that bind tightly and specifically to intrinsically disordered regions (IDRs) of proteins and flexible peptides would have wide application in biological research, therapeutics, and diagnosis. However, the lack of defined structures and the high variability in sequence and conformational preferences has complic...

RhoA is the founding member of the Ras-homology (Rho) small GTPase family and is a master regulator for multiple cell functions. Recurrent RhoA mutations have been identified in several human cancers, particularly in leukemia/lymphoma and gastric cancer. Intriguingly, both gain-of-function and loss-of-function mutations of RhoA are present, suggest...

Biomolecular structure analysis from experimental NMR studies generally relies on restraints derived from a combination of experimental and knowledge-based data. A challenge for the structural biology community has been a lack of standards for representing these restraints, preventing the establishment of uniform methods of model-vs-data structure...

Multidimensional NMR spectra are the basis for studying proteins by NMR spectroscopy and crucial for the development and evaluation of methods for biomolecular NMR data analysis. Nevertheless, in contrast to derived data such as chemical shift assignments in the BMRB and protein structures in the PDB databases, this primary data is in general not p...

Biomolecules exhibit dynamic behavior that single-state models of their structures cannot fully capture. We review some recent advances for investigating multiple conformations of biomolecules, including experimental methods, molecular dynamics simulations, and machine learning. We also address the challenges associated with representing single- an...

Membrane-traversing peptides offer opportunities for targeting intracellular proteins and oral delivery. Despite progress in understanding the mechanisms underlying membrane traversal in natural cell-permeable peptides, there are still several challenges to designing membrane-traversing peptides with diverse shapes and sizes. Conformational flexibi...

Recent advances in molecular modeling of protein structures are changing the field of structural biology. AlphaFold-2 (AF2), an AI system developed by DeepMind, Inc., utilizes attention-based deep learning to predict models of protein structures with high accuracy relative to structures determined by X-ray crystallography and cryo-electron microsco...

Recent advances in molecular modeling of protein structures are changing the field of structural biology. AlphaFold-2 (AF2), an AI system developed by DeepMind, Inc., utilizes attention-based deep learning to predict models of protein structures with high accuracy relative to structures determined by X-ray crystallography and cryo-electron microsco...

We use computational design coupled with experimental characterization to systematically investigate the design principles for macrocycle membrane permeability and oral bioavailability. We designed 184 6–12 residue macrocycles with a wide range of predicted structures containing noncanonical backbone modifications and experimentally determined stru...

NMR is a valuable experimental tool in the structural biologist’s toolkit to elucidate the structures, functions, and motions of biomolecules. The progress of machine learning, particularly in structural biology, reveals the critical importance of large, diverse, and reliable datasets in developing new methods and understanding in structural biolog...

Recent advances in molecular modeling using deep learning have the potential to revolutionize the field of structural biology. In particular, AlphaFold has been observed to provide models of protein structures with accuracies rivaling medium-resolution X-ray crystal structures, and with excellent atomic coordinate matches to experimental protein NM...

Recent advances in molecular modeling using deep learning have the potential to revolutionize the field of structural biology. In particular, AlphaFold has been observed to provide models of protein structures with accuracy rivaling medium-resolution X-ray crystal structures, and with excellent atomic coordinate matches to experimental protein NMR...

NMR is a valuable experimental tool in the structural biologist’s toolkit to elucidate the structures, functions, and motions of biomolecules. The progress of machine learning, particularly in structural biology, reveals the critical importance of large, diverse, and reliable datasets in developing new methods and understanding in structural biolog...

Effective control of COVID-19 requires antivirals directed against SARS-CoV-2. We assessed ten hepatitis C virus (HCV) protease-inhibitor drugs as potential SARS-CoV-2 antivirals. There is a striking structural similarity of the substrate binding clefts of SARS-CoV-2 main protease (Mpro) and HCV NS3/4A protease. Virtual docking experiments show tha...

Effective control of COVID-19 requires antivirals directed against SARS-CoV-2 virus. Here we assess ten available HCV protease inhibitor drugs as potential SARS-CoV-2 antivirals. There is a striking structural similarity of the substrate binding clefts of SARS- CoV-2 M pro and HCV NS3/4A proteases, and virtual docking experiments show that all ten...

Proanthocyanidins (condensed tannins) are important in food chemistry, agriculture, and health, driving demand for improvements in structure determination. We used ultrahigh resolution Fourier transform-ion cyclotron resonance mass spectrometry (FT-ICR MS) methods to determine the exact composition of individual species in heterogeneous mixtures of...

Rational design of protein-polymer bioconjugates is hindered by limited experimental data and mechanistic understanding on interactions between the two. In this communication, nuclear magnetic resonance (NMR) paramagnetic relaxation enhancement (PRE) reports on distances between paramagnetic spin labels and NMR active nuclei, informing on the confo...

SP_0782 from Streptococcus pneumoniae is a dimeric protein that potentially binds with single-stranded DNA (ssDNA) in a manner similar to human PC4, the prototype of PC4-like proteins, which plays roles in transcription and maintenance of genome stability. In a previous NMR study, SP_0782 exhibited an ssDNA-binding property different from YdbC, a p...

Protein CGL2373 from Corynebacterium glutamicum was previously proposed to be a member of the polyketide_cyc2 family, based on amino‐acid sequence and secondary structure features derived from NMR chemical shift assignments. We report here the solution NMR structure of CGL2373, which contains three α‐helices and one antiparallel β‐sheet and adopts...

Growth arrest specific 7 (Gas7) protein is a cytoskeleton regulator playing a crucial role in neural cell development and function, and has been implicated in Alzheimer disease, schizophrenia and cancers. In human, three Gas7 isoforms can be expressed from a single Gas7 gene, while only the longest isoform, hGas7c, possesses an SH3 domain at the N-...

The ever-increasing occurrence of antibiotic resistance presents a major threat to public health. Specifically, resistance conferred by β-lactamases places the efficacy of currently available antibiotics at risk. Klebsiella pneumoniae carbapenemase-2 (KPC-2) is a β-lactamase that enables carbapenem resistance and represents a clear and present dang...

Antibiotic resistance is a continual threat in the clinic. One of the resistance modes bacteria employ is the expression of the beta‐lactamases that target most of the clinically relevant antibiotics, such as penicillin. The enzyme hydrolyzes the β‐lactam bond, which renders the antibiotic ineffective. Targeting β‐lactamases with small molecule inh...

We report the solution NMR structure of CHU_1110 from Cytophaga hutchinsonii. CHU_1110 contains three α‐helices and one antiparallel β‐sheet, forming a large cavity in the center of the protein, which are consistent with the structural characteristics of AHSA1 protein family. This protein shows high structural similarities to the prokaryotic protei...

SH2 domain-containing inositol 5-phosphatase 2 (SHIP2) binds with the Y1356-phosphorylated hepatocyte growth factor (HGF) receptor, c-MET, through its SH2 domain, which is essential for the role of SHIP2 in HGF-induced cell scattering and cell spreading. Previously, the experimental structure of the SH2 domain from SHIP2 (SHIP2-SH2) had not been re...

The forkhead-associated (FHA) domain is known as a phosphopeptide recognition domain embedded in regulatory proteins from both eukaryotes and bacteria with various biological functions. In this study, the gene encoding a predicted FHA domain from Mb1858 (residues V24-D155 from the 162 amino acid protein Mb1858) in Mycobacterium bovis was cloned, an...

Human protein TGIF1 is an essential regulator of cell fate with broad roles in different tissues, and has been implicated in holoprosencephaly (HPE) and many cancers. The function of TGIF1 in transcriptional regulation depends on its three-amino acid loop extension (TALE) type of homeodomain (HD). Two missense mutations that led to P192A and R219C...

Protein CGL2373 from Corynebacterium glutamicum, which is 155 amino acids long and 17.7 kDa, is a member of the polyketide_cyc2 family. As a potential polyketide cyclase, it may play an important role in the biosynthesis of aromatic polyketides that are the source of many bioactive molecules. Here we report the complete (1)H, (13)C and (15)N chemic...

Hsp90 is a homodimeric ATP-dependent molecular chaperone that remodels its substrate ‘client’ proteins, facilitating their folding and activating them for biological function. Despite decades of research, the mechanism connecting ATP hydrolysis and chaperone function remains elusive. Particularly puzzling has been the apparent lack of cooperativity...

Data collection and refinement statistics for zebrafish TRAP1 crystals. Each dataset was collected from a single crystal. Values in parentheses are for highest-resolution shell. DOI: http://dx.doi.org/10.7554/eLife.25235.014

A strategy for acquiring structural information from sparsely isotopically labeled large proteins is illustrated with an application to the E. coli heat-shock protein, HtpG (high temperature protein G), a 145 kDa dimer. It uses (13)C-alanine methyl labeling in a perdeuterated background to take advantage of the sensitivity and resolution of Methyl-...

Aqueous initiators for continuous activator regeneration atom transfer radical polymerization (ICAR ATRP) was applied to graft well defined acrylamide, N,N-dimethylacrylamide and N-vinylimidazole homo and block copolymers from a model protein initiator (bovine serum albumin (BSA)) under bio-relevant conditions. Using N-vinylimidazole as a ligand ca...

Genetic and functional studies showed that some components of the Acinetobacter baumannii ATCC 17978 A1S_0112-A1S_0119 gene cluster are critical for biofilm biogenesis and surface motility. Recently, our group has shown that the A1S_0114 gene was involved in biofilm formation, a process related with pathogenesis. Confirming our previous results, mi...

Ac-505 MS/MS spectra. (A) MS data collected on the FT-ICR with ECD fragmentation in positive ion mode. (B) MS/MS collected on a MaXis QTOF in positive ion mode (LC-MS) and (C) negative ion mode (direct injection).

Confirmation of the polycistronic nature of the A. baumannii ATCC 17978 A1S_0112-A1S_0119 operon. Total DNA (lanes 2–7), cDNA (lanes 8–13) and total RNA (lanes 14–19) samples were used as templates in PCR reactions using primers annealing to the A1S_0112-A1S 0113 (lanes 2, 8, and 14), A1S_0113-A1S_0115 (lanes 3, 9, and 15), A1S_0115-A1S_0116 (lanes...

MS/MS—based fragmentation predictions (A,B) are two of the 40 total MSn spectra from MS(3) to MS(6) recorded on the OSU Bruker amaZon in positive ion mode using direct injection of a HPLC purified Ac-505. MS(2) for 488.3 and MS(4) for 235.2 m/z fragment ions are shown in (A,B), respectively. Neutral losses are shown in black. Panel (C) is a schemat...

Predicted product ions of Ac-505 based on accurate mass and typical bond cleavage patterns under electrospray conditions in positive ion mode.

Predicted product ions of Ac-505 based on accurate mass and typical bond cleavage patterns under electrospray conditions in negative ion mode.

Oligonucleotides used in the present study.

HRMS data of Ac-505 and MS2 fragments measured on a ESI MaXis QTOF (direct infusion) in negative ion mode.

Growth of the 17978 parental and the Δ0114 isogenic deletion derivative strains. The OD600values of each strain grown in LB or SB at 37°C for 24 h with shaking were determined hourly. Error bars represent the standard error (SE) of the mean.

HRMS data of Ac-505. Measured m/z-values are reported for MS fragment ions measured on ESI MaXis QTOF (LC-MS2) and 15T FT-ICR ECD (direct infusion) instruments in positive ion mode, respectively. The molecular formulas of the product ions are based on accurate mass and isotopic pattern matching of the ions as well as typical bond cleavage patterns...

Hsp90 is a homodimeric ATP-dependent molecular chaperone that remodels its substrate " client " proteins, facilitating their folding and activating them for biological function. Despite decades of research, the mechanism connecting ATP hydrolysis and chaperone function remains elusive. Particularly puzzling has been the apparent lack of cooperativi...

We report the solution NMR structure of RHE_CH02687 from Rhizobium etli. Its structure consists of two β-sheets that together with two short and one long α-helix form a hydrophobic cavity. This protein shows a high structural similarity to the prokaryotic protein YndB from Bacillus subtilis, and the eukaryotic protein Aha1. NMR titration experiment...

Human INSM1 containing five C-terminal C2H2-type zinc fingers (ZFs), is a key regulator of neuroendocrine development. Previous research reported that full-length INSM1 containing all five ZFs recognized a consensus DNA sequence. Structure elucidation of human INSM1 ZFs is currently insufficient to understand the DNA binding mechanism. Herein, we p...

Importance: In this study, we found that our P[19] strain and other P[II] RVs recognize mucin cores and the type 1 HBGA precursors as the minimal functional units and that additional saccharides adjacent to these units can alter binding outcomes and thereby possibly host ranges. These data may help to explain why some P[II] RVs, such as P[6] and P...

The protein SP_0782 from Streptococcus pneumonia is a small homodimeric protein that belongs to a protein family containing representative members with single-stranded DNA (ssDNA) binding functions. The ssDNA binding of the homolog YdbC from Lactococcus lactis was previously characterized when bound to a 20-mer of pyridine-rich ssDNA, sharing an ov...

We have developed an on-line NMR/X-ray Structure Pair Data Repository. The NIGMS Protein Structure Initiative (PSI) has provided many valuable reagents, 3D structures, and technologies for structural biology. The Northeast Structural Genomics Consortium was one of several PSI centers. NESG used both X-ray crystallography and NMR spectroscopy for pr...

The success of homodimer structure determination by conventional solution NMR spectroscopy relies greatly on interchain distance restraints (less than 6 Å) derived from nuclear Overhauser effects (NOEs) obtained from (13)C-edited, (12)C-filtered NOESY experiments. However, these experiments may fail when the mixed (13)C-/(12)C-homodimer is never si...

Calicheamicin ɣ1I (1) is an enediyne antitumor compound produced by Micromonospora echinospora spp. calichensis and its biosynthetic gene cluster has been previously reported. Despite extensive analysis and biochemical study, several genes in the biosynthetic gene cluster of 1 remain functionally unassigned. Using a structural genomics approach and...

We have determined the solution NMR structure of the intermembrane space domain (IMSD) of the human mitochondrial ATPase associated with various activities (AAA) protease known as AFG3-like protein 2 (AFG3L2). Our structural analysis and molecular dynamics results indicate that the IMSD is peripherally bound to the membrane surface. This is a modif...

CV_2116 is a small hypothetical protein of 82 amino acids from the Gram-negative coccobacillus Chromobacterium violaceum. A PSI-BLAST search using the CV_2116 sequence as a query identified only one hit (E = 2e(-07)) corresponding to a hypothetical protein OR16_04617 from Cupriavidus basilensis OR16, which failed to provide insight into the functio...

Here, we report novel methods to measure rate constants for homodimer subunit exchange using double electron–electron resonance (DEER) electron paramagnetic resonance spectroscopy measurements and nuclear magnetic resonance spectroscopy based paramagnetic relaxation enhancement (PRE) measurements. The techniques were demonstrated using the homodime...

The solution nuclear magnetic resonance (NMR) structures and backbone (15)N dynamics of the specialized acyl carrier protein (ACP), RpAcpXL, from Rhodopseudomonas palustris, in both the apo form and holo form modified by covalent attachment of 4'-phosphopantetheine at S37, are virtually identical, monomeric, and correspond to the closed conformatio...

The protein Pspto_3016 is a 117-residue member of the protein domain family PF04237 (DUF419), which is to date a functionally uncharacterized family of proteins. In this report, we describe the structure of Pspto_3016 from Pseudomonas syringae solved by both solution NMR and X-ray crystallography at 2.5 Å resolution. In both cases, the structure of...

The protein domain family PF12095 (DUF3571) is a functionally uncharacterized family of small proteins conserved from cyanobacteria to plants that are typically 85 to 95 amino acids in length in cyanobacteria. In this report, we describe the solution NMR structure of the 86-residue protein Asl3597 from Nostoc sp. PCC7120. The structure of Asl3597,...