Amino Acids, Peptides, and Proteins Commons^™

Dual Engagement Of The Nlrp3 And Aim2 Inflammasomes By Plasmodium-Derived Hemozoin And Dna During Malaria, Parisa Kalantari, Rosane B. Deoliveira, Jennie Chan, Yolanda Corbett, Vijay A. K. Rathinam, Andrea Stutz, Eicke Latz, Ricardo T. Gazzinelli, Douglas T. Golenbock, Katherine A. Fitzgerald

Katherine A. Fitzgerald

Hemozoin (Hz) is the crystalline detoxification product of hemoglobin in Plasmodium-infected erythrocytes. We previously proposed that Hz can carry plasmodial DNA into a subcellular compartment that is accessible to Toll-like receptor 9 (TLR9), inducing an inflammatory signal. Hz also activates the NLRP3 inflammasome in primed cells. We found that Hz appears to colocalize with DNA in infected erythrocytes, even before RBC rupture or phagolysosomal digestion. Using synthetic Hz coated in vitro with plasmodial genomic DNA (gDNA) or CpG oligodeoxynucleotides, we observed that DNA-complexed Hz induced TLR9 translocation, providing a priming and an activation signal for inflammasomes. After phagocytosis, Hz and …

A Microrna-1280/Jag2 Network Comprises A Novel Biological Target In High-Risk Medulloblastoma, Fengfei Wang, Marc Remke, Tze-Chen Hsieh, Lizi Wu, Cynthia Hawkins, Joseph M. Wu, Erxi Wu

NYMC Faculty Publications

Over-expression of PDGF receptors (PDGFRs) has been previously implicated in high-risk medulloblastoma (MB) pathogenesis. However, the exact biological functions of PDGFRα and PDGFRβ signaling in MB biology remain poorly understood. Here, we report the subgroup specific expression of PDGFRα and PDGFRβ and their associated biological pathways in MB tumors. c-MYC, a downstream target of PDGFRβ but not PDGFRα, is involved in PDGFRβ signaling associated with cell proliferation, cell death, and invasion. Concurrent inhibition of PDGFRβ and c-MYC blocks MB cell proliferation and migration synergistically. Integrated analysis of miRNA and miRNA targets regulated by both PDGFRβ and c-MYC reveals that increased …

Effect Of Hydrogen Peroxide On The Biosynthesis Of Heme And Proteins: Potential Implications For The Partitioning Of Glu-TrnaGlu Between These Pathways, Carolina Farah, Gloria Levicán, Michael Ibba, Omar Orellana

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Glutamyl-tRNA (Glu-tRNA^Glu) is the common substrate for both protein translation and heme biosynthesis via the C5 pathway. Under normal conditions, an adequate supply of this aminoacyl-tRNA is available to both pathways. However, under certain circumstances, Glu-tRNA^Glu can become scarce, resulting in competition between the two pathways for this aminoacyl-tRNA. In Acidithiobacillus ferrooxidans, glutamyl-tRNA synthetase 1 (GluRS1) is the main enzyme that synthesizes Glu-tRNA^Glu. Previous studies have shown that GluRS1 is inactivated in vitro by hydrogen peroxide (H₂O₂). This raises the question as to whether H₂O₂ negatively affects …

Strengths And Weaknesses Of Hybrid Tpr Technology For Obtaining Structural And Mechanistic Insights Into Tpr Proteins, Shanshan Yu

Theses and Dissertations (ETD)

Tetratricopeptide (TPR) repeats are a 34-residue helix-turn-helix motif that when repeated pack into a superhelical structure. TPR domains are frequently found mediating protein-protein interactions, often through a central groove. One protein complex bearing numerous TPR repeats is the Anaphase Promoting Complex (APC). The anaphase-promoting complex (APC) is a multi-subunit complex, which orchestrates mitotic cell cycles. APC is an E3 ligase in the ubiquitin cascade, and directs the 26S proteosome degradation of cell-cycle regulators. Throughout mitotic progression, proteins that are key regulators of the cell cycle are assembled with polyubiquitin chains by APC.

One domain of the human APC is comprised …

Tuning Responsiveness Of Polypeptide Based Block Copolymers For Drug Delivery, Ashley J. Johnson

Dissertations

The goal of this dissertation was to tune the pH response and self-assembled morphologies of amphiphilic polypeptide block copolymers for use as drug delivery vehicles. Poly(L-lysine) and poly(L-glutamtic acid) are responsive, ionizable polypeptides that undergo secondary structure transitions, from α-helix to random coil, whereby the change in conformation of the peptide chain results in changes to the global morphology of a self-assembled system. The main focus of this work was to understand how changes in the polymer composition and the local environment can lead to control over the behavior of the overall system. First, the responsive behavior of poly(L-lysine) block …

Synchronous Opening And Closing Motions Are Essential For Camp-Dependent Protein Kinase A Signaling, Atul K. Srivastava, Leanna R. Mcdonald, Alessandro Cembran, Jonggul Kim, Larry R. Masterson, Christopher L. Mcclendon, Susan S. Taylor, Gianluigi Veglia

Larry Masterson

Regulated Splicing Of The Alpha6 Integrin Cytoplasmic Domain Determines The Fate Of Breast Cancer Stem Cells, Hira Lal Goel, Tatiana Gritsko, Bryan Pursell, Cheng Chang, Leonard D. Shultz, Dale L. Greiner, Jens Henrik Norum, Rune Toftgard, Leslie M. Shaw, Arthur M. Mercurio

Arthur M. Mercurio

Although the alpha6beta1 integrin has been implicated in the function of breast and other cancer stem cells (CSCs), little is known about its regulation and relationship to mechanisms involved in the genesis of CSCs. We report that a CD44(high)/CD24(low) population, enriched for CSCs, is comprised of distinct epithelial and mesenchymal populations that differ in expression of the two alpha6 cytoplasmic domain splice variants: alpha6A and alpha6B. alpha6Bbeta1 expression defines the mesenchymal population and is necessary for CSC function, a function that cannot be executed by alpha6A integrins. The generation of alpha6Bbeta1 is tightly controlled and occurs as a consequence of …

Peptide Arrays For Detecting Naphthenic Acids In Oil Sands Process Affected Water, Kamaljit Kaur, Subir Bhattacharjee, Rajesh G. Pillai, Sahar Ahmed, Sarfuddin Azmi

Pharmacy Faculty Articles and Research

Naphthenic acids (NAs) are water-soluble components of petroleum. The characterization and quantification of NAs by analytical methods have proved quite challenging, whilst the toxic effects of these water-soluble compounds on a variety of organisms adversely affecting reproduction and steroid production is becoming apparent. In this study, we report a fluorescence-based competitive binding method for rapid sensing of the presence of NAs using cellulosic peptide array strips as sensors. The peptide array was designed from sequences derived from the estrogen receptor (ER). Several of these peptides were able to detect the presence of NAs at low micromolar (∼5 mg L^{−1 …}

The Translational Biology Of Remyelination: Past, Present, And Future, Robin J.M. Franklin, Vittorio Gallo

Pediatrics Faculty Publications

Amongst neurological diseases, multiple sclerosis (MS) presents an attractive target for regenerative medicine. This is because the primary pathology, the loss of myelin-forming oligodendrocytes, can be followed by a spontaneous and efficient regenerative process called remyelination. While cell transplantation approaches have been explored as a means of replacing lost oligodendrocytes, more recently therapeutic approaches that target the endogenous regenerative process have been favored. This is in large part due to our increasing understanding of (1) the cell types within the adult brain that are able to generate new oligodendrocytes, (2) the mechanisms and pathways by which this achieved, and (3) …

Hdac8 And Stat3 Repress Bmf Gene Activity In Colon Cancer Cells, Y Kang, Hui Nian, P Rajendran, W Dashwood, John T. Pinto, E Ho, R Dashwood

NYMC Faculty Publications

Histone deacetylase (HDAC) inhibitors are undergoing clinical trials as anticancer agents, but some exhibit resistance mechanisms linked to anti-apoptotic Bcl-2 functions, such as BH3-only protein silencing. HDAC inhibitors that reactivate BH3-only family members might offer an improved therapeutic approach. We show here that a novel seleno-α-keto acid triggers global histone acetylation in human colon cancer cells and activates apoptosis in a p21-independent manner. Profiling of multiple survival factors identified a critical role for the BH3-only member Bcl-2-modifying factor (Bmf). On the corresponding BMF gene promoter, loss of HDAC8 was associated with signal transducer and activator of transcription 3 (STAT3)/specificity protein …

Targeting The Small- And Intermediate Conductance Ca2+- Activated Potassium Channels: The Drug Binding Pocket At The Channel/Calmodulin Interface, Meng Cui, Guangrong Qin, Kunqian Yu, M. Scott Bowers, Miao Zhang

Pharmacy Faculty Articles and Research

The small- and intermediate-conductance Ca 2+ -activated potassium (SK/IK) channels play important roles in the regulation of excitable cells in both the central nervous and cardiovascular systems. Evidence from animal models has implicated SK/IK channels in neurological conditions such as ataxia and alcohol use disorders. Further, genome-wide association studies have suggested that cardiovascular abnormalities such as arrhythmias and hypertension are associated with single nucleotide polymorphisms that occur within the genes encoding the SK/IK channels. The Ca 2+ sensitivity of the SK/IK channels stems from a constitutively bound Ca 2+ -binding protein: calmodulin. Small-molecule positive modulators of SK/IK channels have been …

The Non-Canonical Hydroxylase Structure Of Yfcm Reveals A Metal Ion-Coordination Motif Required For Ef-P Hydroxylation, Kan Kobayashi, Assaf Katz, Andrei Rajkovic, Ryohei Ishii, Owen E. Branson, Michael A. Freitas, Ryuichiro Ishitani, Michael Ibba, Osamu Nureki

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

EF-P is a bacterial tRNA-mimic protein, which accelerates the ribosome-catalyzed polymerization of poly-prolines. In Escherichia coli, EF-P is post-translationally modified on a conserved lysine residue. The post-translational modification is performed in a two-step reaction involving the addition of a β-lysine moiety and the subsequent hydroxylation, catalyzed by PoxA and YfcM, respectively. The β-lysine moiety was previously shown to enhance the rate of poly-proline synthesis, but the role of the hydroxylation is poorly understood. We solved the crystal structure of YfcM and performed functional analyses to determine the hydroxylation mechanism. In addition, YfcM appears to be structurally distinct from any …

Mistranslation Of The Genetic Code, Adil Moghal, Kyle Mohler, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

During mRNA decoding at the ribosome, deviations from stringent codon identity, or “mistranslation,” are generally deleterious and infrequent. Observations of organisms that decode some codons ambiguously, and the discovery of a compensatory increase in mistranslation frequency to combat environmental stress have changed the way we view “errors” in decoding. Modern tools for the study of the frequency and phenotypic effects of mistranslation can provide quantitative and sensitive measurements of decoding errors that were previously inaccessible. Mistranslation with non‐protein amino acids, in particular, is an enticing prospect for new drug therapies and the study of molecular evolution.

Relaxed Substrate Specificity Leads To Extensive Trna Mischarging By Streptococcus Pneumoniae Class I And Class Ii Aminoacyl-Trna Synthetases, Jennifer Shepherd, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Aminoacyl-tRNA synthetases provide the first step in protein synthesis quality control by discriminating cognate from noncognate amino acid and tRNA substrates. While substrate specificity is enhanced in many instances by cis- and trans-editing pathways, it has been revealed that in organisms such as Streptococcus pneumoniae some aminoacyl-tRNA synthetases display significant tRNA mischarging activity. To investigate the extent of tRNA mischarging in this pathogen, the aminoacylation profiles of class I isoleucyl-tRNA synthetase (IleRS) and class II lysyl-tRNA synthetase (LysRS) were determined. Pneumococcal IleRS mischarged tRNA^Ile with both Val, as demonstrated in other bacteria, and Leu in a tRNA sequence-dependent …

Selective Phosphorylation Modulates The Pip2 Sensitivity Of The Cam-Sk Channel Complex, Miao Zhang, Xuan-Yu Meng, Meng Cui, John M. Pascal, Diomedes E. Logothetis, Ji-Fang Zhang

Pharmacy Faculty Articles and Research

Phosphatidylinositol bisphosphate (PIP2) regulates the activities of many membrane proteins including ion channels through direct interactions. However, the affinity of PIP2 is so high for some channel proteins that its physiological role as a modulator has been questioned. Here we show that PIP2 is an important cofactor for activation of small conductance Ca2+-activated potassium channels (SK) by Ca2+-bound calmodulin (CaM). Removal of the endogenous PIP2 inhibits SK channels. The PIP2-binding site resides at the interface of CaM and the SK C-terminus. We further demonstrate that the affinity of PIP2 for its target proteins can be regulated by cellular signaling. Phosphorylation …

Ligand-Receptor Interactions For Supramolecular Disassembly With Applications In Screening And Drug Delivery, Diego Amado Torres

Doctoral Dissertations

Proteins have the capacity to bind specific sets of compounds known as ligands, these are small molecules with a recurrent theme in their molecular design that is a characteristic exploited here to (i) identify particular affinities of small molecules for proteins with the aim of using them as ligands, inhibitors, or targeting moieties in more complex systems by means of a methodology that screens small molecules based on protein affinity; (ii) decorate a self-assembling supramolecular system at different positions, making it responsive to a complementary protein with the aim of exploring differences in disassembly and sensitivity of the release of …

Translation Initiation Rate Determines The Impact Of Ribosome Stalling On Bacterial Protein Synthesis, Steven J. Hersch, Sara Elgamal, Assaf Katz, Michael Ibba, William Wiley Navarre

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Ribosome stalling during translation can be caused by a number of characterized mechanisms. However, the impact of elongation stalls on protein levels is variable, and the reasons for this are often unclear. To investigate this relationship, we examined the bacterial translation elongation factor P (EF-P), which plays a critical role in rescuing ribosomes stalled at specific amino acid sequences including polyproline motifs. In previous proteomic analyses of both Salmonella and Escherichia coli efp mutants, it was evident that not all proteins containing a polyproline motif were dependent on EF-P for efficient expression in vivo . The α- and β-subunits of …

Ef-P Dependent Pauses Integrate Proximal And Distal Signals During Translation, Sara Elgamal, Assaf Katz, Steven J. Hersch, David Newsom, Peter White, William Wiley Navarre, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Elongation factor P (EF-P) is required for the efficient synthesis of proteins with stretches of consecutive prolines and other motifs that would otherwise lead to ribosome pausing. However, previous reports also demonstrated that levels of most diprolyl-containing proteins are not altered by the deletion of efp. To define the particular sequences that trigger ribosome stalling at diprolyl (PPX) motifs, we used ribosome profiling to monitor global ribosome occupancy in Escherichia coli strains lacking EF-P. Only 2.8% of PPX motifs caused significant ribosomal pausing in the Δefp strain, with up to a 45-fold increase in ribosome density observed at …

Sma-Causing Missense Mutations In Survival Motor Neuron (Smn) Display A Wide Range Of Phenotypes When Modeled In Drosophila, Kavita Praveen, Ying Wen, Kelsey M. Gray, John J. Noto, Akash R. Patlolla, Gregory D. Van Duyne, A. Gregory Matera

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Mutations in the human survival motor neuron 1 (SMN) gene are the primary cause of spinal muscular atrophy (SMA), a devastating neuromuscular disorder. SMN protein has a well-characterized role in the biogenesis of small nuclear ribonucleoproteins (snRNPs), core components of the spliceosome. Additional tissue-specific and global functions have been ascribed to SMN; however, their relevance to SMA pathology is poorly understood and controversial. Using Drosophila as a model system, we created an allelic series of twelve Smn missense mutations, originally identified in human SMA patients. We show that animals expressing these SMA-causing mutations display a broad range of …

Submonomer Synthesis And Structure-Activity Relationship Studies Of Azapeptide Inhibitors Of The Insulin Receptor Tyrosine Kinase, Lathamol A. Kurian

Seton Hall University Dissertations and Theses (ETDs)

Azapeptides are a class of peptide mimics (peptidomimetics), which have served as valuable tools for the development of peptide based therapeutic agents. The therapeutic promise of azapeptides has been correlated to its primary sequence modification which translates into bio-active secondary structures that improves the pharmacological properties of the native peptide sequence. More specifically, azapeptides contain a semicarbazide within the peptide backbone which restricts the peptide bond torsion angles (φ, ψ) into pre-organized b-turn secondary structures. Thus, azapeptides have been shown to stabilize bio-active b-turn secondary structures responsible for high affinity and selective binding to a target …

Mapping The Human Vasculature By In Vivo Phage Display, Julianna Bronk

Dissertations & Theses (Open Access)

In vivo phage display screenings by intravenous injection of a random phage-displayed peptide library allow for the selection of peptides that localize to specific vascular beds. At the University of Texas MD Anderson Cancer Center, we have had the opportunity to perform phage display screenings in cancer patients in order to select for cancer specific targets directly in humans. These targets serve to define biochemical diversity of endothelial cell surfaces and can be validated and explored towards the design of vascular-targeted pharmacology. In the most recent patient screen, samples were recovered from hepatocellular carcinoma (HCC) as well as 26 additional …

Trnas As Regulators Of Biological Processes, Medha Raina, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Transfer RNAs (tRNA) are best known for their role as adaptors during translation of the genetic code. Beyond their canonical role during protein biosynthesis, tRNAs also perform additional functions in both prokaryotes and eukaryotes for example in regulating gene expression. Aminoacylated tRNAs have also been implicated as substrates for non-ribosomal peptide bond formation, post-translational protein labeling, modification of phospholipids in the cell membrane, and antibiotic biosyntheses. Most recently tRNA fragments, or tRFs, have also been recognized to play regulatory roles. Here, we examine in more detail some of the new functions emerging for tRNA in a variety of cellular processes …

Oxidation Of Cellular Amino Acid Pools Leads To Cytotoxic Mistranslation Of The Genetic Code, Tammy J. Bullwinkle, Noah M. Reynolds, Medha Raina, Adil Moghal, Eleftheria Matsa, Andrei Rajkovic, Huseyin Kayadibi, Farbod Fazlollahi, Christopher Ryan, Nathaniel Howitz, Kym F. Faull, Beth A. Lazazzera, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Aminoacyl-tRNA synthetases use a variety of mechanisms to ensure fidelity of the genetic code and ultimately select the correct amino acids to be used in protein synthesis. The physiological necessity of these quality control mechanisms in different environments remains unclear, as the cost vs benefit of accurate protein synthesis is difficult to predict. We show that in Escherichia coli, a non-coded amino acid produced through oxidative damage is a significant threat to the accuracy of protein synthesis and must be cleared by phenylalanine-tRNA synthetase in order to prevent cellular toxicity caused by mis-synthesized proteins. These findings demonstrate how stress …

Endothelial And Smooth Muscle-Dependent Vascular Reactivity In Immature Arterialized Collateral Capillaries, Caitlin Koeroghlian

Biomedical Engineering

Peripheral arterial occlusive disease (PAOD) occurs due to the build up of atherosclerotic plaque and reduces blood flow to cause chronic ischemia. Patients with PAOD may experience intermittent claudication, or the pain in limb skeletal muscles due to a decease in blood flow. Collateral arteries can act as a natural bypass and improve blood flow to hypoxic tissue by creating an alternate route for blood to flow, but not all patients with PAOD have pre-existing collateral networks. Animal studies indicate that tissues without pre-existing collateral networks can form de novo collaterals from capillaries following occlusion of a feed artery. Unfortunately, …

Structural Similarity Between Β3-Peptides Synthesized From Β3-Homo-Amino Acids Or L-Aspartic Acid Monomers, Sahar Ahmed, Tara Sprules, Kamaljit Kaur

Pharmacy Faculty Articles and Research

Formation of stable secondary structures by oligomers that mimic natural peptides is a key asset for enhanced biological response. Here we show that oligomeric β³‐hexapeptides synthesized from l‐aspartic acid monomers (β³‐peptides 1, 5a, and 6) or homologated β³‐amino acids (β³‐peptide 2), fold into similar stable 14‐helical secondary structures in solution, except that the former form right‐handed 14‐helix and the later form left‐handed 14‐helix. β³‐Peptides from l‐Asp monomers contain an additional amide bond in the side chains that provides opportunities for more hydrogen bonding. However, based on the …

Reduced Amino Acid Specificity Of Mammalian Tyrosyl-Trna Synthetase Is Associated With Elevated Mistranslation Of Tyr Codons, Medha Raina, Adil Moghal, Amanda Kano, Mathew Jerums, Paul D. Schnier, Shun Luo, Rohini Deshpande, Pavel D. Bondarenko, Henry Lin, Michael Ibba

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Quality control operates at different steps in translation to limit errors to approximately one mistranslated codon per 10,000 codons during mRNA-directed protein synthesis. Recent studies have suggested that error rates may actually vary considerably during translation under different growth conditions. Here we examined the misincorporation of Phe at Tyr codons during synthesis of a recombinant antibody produced in tyrosine-limited Chinese hamster ovary (CHO) cells. Tyr to Phe replacements were previously found to occur throughout the antibody at a rate of up to 0.7% irrespective of the identity or context of the Tyr codon translated. Despite this comparatively high mistranslation rate, …

Syntheses Of Precursors To Fluorine-18 Labeled Pet Imaging Agents, Lindsay B. Boling

Chancellor’s Honors Program Projects

No abstract provided.

Phage Display Library Screening For Psa-/Lo Prostate Cancer Cell-Binding Peptides, John R. Moore

Dissertations & Theses (Open Access)

Prostate cancer (PCa) is one of the leading malignancies affecting men worldwide. Our lab focuses on understanding the molecular mechanisms underlying prostate carcinogenesis and developing therapeutics that target the cells responsible for driving PCa and mediating therapy resistance. My master thesis research employs a phage display library screening technology aiming to identify peptides that preferentially home in to undifferentiated PCa cells, which our lab has previously demonstrated to be intrinsically resistant to castration.

There is now evidence that a population of cells in PCa possesses characteristics associated with stem cells; these cells are referred to as cancer stem cells (CSCs). …

Analysis Of Ligand Bias In Functional Studies Involving The Allosteric Modulation Of G Protein- Coupled Receptors, Frederick J. Ehlert, Michael T. Griffin

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Introduction

The affinity constants of a ligand for active and inactive states of a receptor ultimately determine its capacity to activate downstream signaling events. In this report, we describe a reverse-engineering strategy for estimating these microscopic constants.

Methods

Our approach involves analyzing responses measured downstream in the signaling pathway of a G protein-coupled receptor under conditions of allosteric modulation and reduced receptor expression or partial receptor inactivation. The analysis also yields estimates of the isomerization constant of the unoccupied receptor, the sensitivity constant of the signaling pathway, and the more empirical parameters of the receptor population including the observed affinities …

Anti-Gd2 Etoposide-Loaded Immunoliposomes For The Treatment Of Gd2 Positive Tumors, Brandon S. Brown

Dissertations & Theses (Open Access)

Systemic chemotherapeutics remain the standard of care for most malignancies even though they frequently suffer from narrow therapeutic index, poor serum solubility, and off-target effects. Monoclonal antibodies that specifically bind antigens overexpressed on many tumors such as the ganglioside, GD2, can be conjugated to drug-loaded liposomes to create a targeted drug delivery system. In this study, we have encapsulated etoposide, a topoisomerase inhibitor effective against a wide range of cancers, in surface modified liposomes decorated with anti-GD2 antibodies. We characterized the properties of the liposomes using a variety of methods including dynamic light scattering, electron microscopy, and Fourier transformed infrared …