Mutations at the Subunit Interface of Yeast Proliferating Cell Nuclear Antigen Reveal a Versatile Regulatory Domain

Acknowledgments We thank Szilvia Minorits for technical assistance. I.U. conceived and designed the project and wrote the manuscript. All authors participated in designing and performing the experiments, and analyzing the results. The authors declare no competing financial interests. This work was also supported by a grant from the National Research, Development and Innovation Office GINOP-2.3.2-15-2016-00001. Funding: This work was supported by Hungarian Science Foundation Grant OTKA 109521 and National Research Development and Innovation Office GINOP-2.3.2-15-2016-00001. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Peer reviewedPublisher PD

DNA mimicry by a high-affinity anti-NF-κB RNA aptamer

The binding of RNA molecules to proteins or other ligands can require extensive RNA folding to create an induced fit. Understanding the generality of this principle involves comparing structures of RNA before and after complex formation. Here we report the NMR solution structure of a 29-nt RNA aptamer whose crystal structure had previously been determined in complex with its transcription factor target, the p502 form of NF-κB. The RNA aptamer internal loop structure has pre-organized features that are also found in the complex, including non-canonical base pairing and cross-strand base stacking. Remarkably, the free RNA aptamer structure possesses a major groove that more closely resembles B-form DNA than RNA. Upon protein binding, changes in RNA structure include the kinking of the internal loop and distortion of the terminal tetraloop. Thus, complex formation involves both pre-formed and induced fit binding interactions. The high affinity of the NF-κB transcription factor for this RNA aptamer may largely be due to the structural pre-organization of the RNA that results in its ability to mimic DNA

Global Conformational Dynamics of a Y-Family DNA Polymerase during Catalysis

High-resolution analysis of protein, and DNA conformational changes during DNA polymerization, established relationships between the enzymatic function and conformational dynamics of individual domains for a DNA polymerase

The RAGNYA fold: a novel fold with multiple topological variants found in functionally diverse nucleic acid, nucleotide and peptide-binding proteins

Using sensitive structure similarity searches, we identify a shared α+β fold, RAGNYA, principally involved in nucleic acid, nucleotide or peptide interactions in a diverse group of proteins. These include the Ribosomal proteins L3 and L1, ATP-grasp modules, the GYF domain, DNA-recombination proteins of the NinB family from caudate bacteriophages, the C-terminal DNA-interacting domain of the Y-family DNA polymerases, the uncharacterized enzyme AMMECR1, the siRNA silencing repressor of tombusviruses, tRNA Wybutosine biosynthesis enzyme Tyw3p, DNA/RNA ligases and related nucleotidyltransferases and the Enhancer of rudimentary proteins. This fold exhibits three distinct circularly permuted versions and is composed of an internal repeat of a unit with two-strands and a helix. We show that despite considerable structural diversity in the fold, its representatives show a common mode of nucleic acid or nucleotide interaction via the exposed face of the sheet. Using this information and sensitive profile-based sequence searches: (1) we predict the active site, and mode of substrate interaction of the Wybutosine biosynthesis enzyme, Tyw3p, and a potential catalytic role for AMMECR1. (2) We provide insights regarding the mode of nucleic acid interaction of the NinB proteins, and the evolution of the active site of classical ATP-grasp enzymes and DNA/RNA ligases. (3) We also present evidence for a bacterial origin of the GYF domain and propose how this version of the fold might have been utilized in peptide interactions in the context of nucleoprotein complexes

How glutaminyl-tRNA synthetase selects glutamine

AbstractBackground: Aminoacyl-tRNA synthetases covalently link a specific amino acid to the correct tRNA. The fidelity of this reaction is essential for accurate protein synthesis. Each synthetase has a specific molecular mechanism to distinguish the correct pair of substrates from the pool of amino acids and isologous tRNA molecules. In the case of glutaminyl-tRNA synthetase (GlnRS) the prior binding of tRNA is required for activation of glutamine by ATP. A complete understanding of amino acid specificity in GlnRS requires the determination of the structure of the synthetase with both tRNA and substrates bound.Results: A stable glutaminly-adenylate analog, which inhibits GlnRS with a Ki of 1.32 μM, was synthesized and cocrystallized with GlnRS and tRNA2Gln. The crystal structure of this ternary complex has been refined at 2.4 å resolution and shows the interactions made between glutamine and its binding site.Conclusions: To select against glutamic acid or glutamate, both hydrogen atoms of the nitrogen of the glutamine sidechain are recognized. The hydroxyl group of Tyr211 and a water molecule are responsible for this recognition; both are obligate hydrogen-bond acceptors due to a network of interacting sidechains and water molecules. The prior binding of tRNAGln that is required for amino acid activation may result from the terminal nucleotide, A76, packing against and orienting Tyr211, which forms part of the amino acid binding site

Formation of virus-like clusters is an intrinsic property of the tumor necrosis factor family member BAFF (B cell activating factor).

The oligomeric state of BAFF (B cell activing factor), a tumor necrosis factor (TNF) family cytokine that plays a critical role in B cell development and survival, has been the subject of recent debate. Myc-tagged BAFF starting at residue Gln136 was previously reported to crystallize as trimers at pH 4.5, whereas a histidine-tagged construct of BAFF, starting at residue Ala134, formed a virus-like cluster containing 60 monomers when crystallized at pH 9.0. The formation of the BAFF 60-mer was pH dependent, requiring pH >or= 7.0. More recently, 60-mer formation was suggested to be artificially induced by the histidine tag, and it was proposed that BAFF, like all other TNF family members, is trimeric. We report here that a construct of BAFF with no amino-terminal tag (Ala134-BAFF) can form a 60-mer in solution. Using size exclusion chromatography and static light scattering to monitor trimer to 60-mer ratios in BAFF preparations, we find that 60-mer formation is pH-dependent and requires histidine 218 within the DE loop of BAFF. Biacore measurements established that the affinity of Ala134-BAFF for the BAFF receptor BAFFR/BR3 is similar to that of myc-Gln136-BAFF, which is exclusively trimeric in solution. However, Ala134-BAFF is more efficacious than myc-Gln136-BAFF in inducing B cell proliferation in vitro. We additionally show that BAFF that is processed and secreted by 293T cells transfected with full-length BAFF, or by a histiocytic lymphoma cell line (U937) that expresses BAFF endogenously, forms a pH-dependent 60-mer in solution. Our results indicate that the formation of the 60-mer in solution by the BAFF extracellular domain is an intrinsic property of the protein, and therefore that this more active form of BAFF may be physiologically relevant

The anti-NgR1 antibody, 1D9, rescues rat retinal ganglion cells after optic nerve transection and ocular hypertension-induced glaucoma

The neuronal leucine rich repeat protein, Nogo66 receptor [NgR1], interacts with at least three CNS myelin proteins [Nogo, MAG and OMgp] and mediates the inhibition of neurite growth. Here we report a monoclonal anti-NgR1 antibody, 1D9, that in addition to inhibiting these interactions in vitro, exhibits neuroprotective properties in vitro and in vivo. Structural analyses performed on the co-crystal complex of the 1D9 Fab and a soluble fragment of NgR1 (sNgR310) indicate that this antibody binds near the junction of the N-terminus cap and leucine rich repeat domain on NgR1. Treatment with 1D9 protected primary neuronal cultures from insults derived from serum withdrawal. Direct intravitreal administration of 1D9 Fab, but not the full 1D9 mAb, consistently promoted the survival of retinal ganglion cells in an optic nerve transection model and an ocular hypertension induced glaucoma model in rat. The lack of activity of the full 1D9 mAb may be partially attributed to NgR1 activation via receptor cross-linking as demonstrated by rhoA activation assay. These results suggest that 1D9 Fab may confer neuroprotection to specific subsets of neurons. Equal contribution: B Hu, A Jirik, and Q Fu Corresponding authors: KF So and DHS Le

Dental Education Resources - historical

Walker, Paul O; Vukovich A.; Berendt, [], Scenes depicting attitudes of a clinical faculty member and a dental hygiene student and how they affect each other. Used for in-service program for dental hygiene faculty. (2:21) Orig. air date: MAY 26 72Walker, Paul O; Vukovich A.; Berendt, [], Scene depicting a dental hygiene student in a clinical experience; poor use of appointment time. Used for in-service program for Dental Hygiene faculty. (2:06) Orig. air date: MAY 26 72Walker, Paul O; Vukovich A.; Berendt, [], Enactment ofinteraction between adental hygiene clinical student and a new instructor; typical problems in this situation are demonstrated. (1:23) Orig. air date: MAY 26 72Robinson, Barbara, "Skills in Understanding the Other Person": This tape demonstrates two communication skills, paraphrasing and perceptions check to help the dental professional understand other people better. (8:01) Orig. air date: JUN 3 74O'Connor, Patricia; Silvian, Joseph, "Trigger Tapes - Workshop": Seven student-faculty lab or clinic situations are presented. The tapes are designed to trigger discussion of faculty members' behavior in the given situations. (15:02) Orig. air date: OCT 23 73Fisch, A. Linc, "Project 119 - Trigger Tapes on Learning/Teaching, Part I": Designed to generate discussion about clinic ethics. Several role playing situations are presented. (26:00) Orig. air date: FEB 9 72Fisch, A. Linc, "Project 119 - Trigger Tapes on Learning/Teaching, Part II": Presents staged situations that trigger""a dialogue on"" different aspects and problems in the teaching of Dental Students. (27:48) Orig. air date: FEB 9 72Walker, Paul O; Vukovich A.; Berendt, [], "Inconsistency": Deals with the problems students have when two different instructors review their work. (2:25) Orig. air date: MAR 1 72Comstock, Frank, "Dental Cubicle, Pt. 1- The Dental Unit": Introduces the University of Michigan School of Dentistry cubicle to the student. Shows the operation of the equipment. (28:07) Orig. air date: SEP 18 72Bagramian, Robert; Strawn, [], "Patient Education for Dental Health": Demonstrates a four step patient education sequence in use in private practices. (27:15) Orig. air date: OCT 8 75Bagramian, Robert, "Patient Education: Asset or Liability": Three successful practitioners discuss the role of patient education in Dentistry. The benefits, as well as the drawbacks, to the practitioner and the patient are covered. (19:32) Orig. air date: JAN 31 76Pape, Bud; Souers, [], "Your Role In Oral Hygiene": A patient education tape showing oral self exam, effects of dental plaque and way of dealing with plaque, use of disclosing solution, tooth brushing, and flossing. (19:21) Orig. air date: NOV 16 76Miller, Doris, "Trigger Tape - Patient Education": Five simulations depict interactions between a patient and Hygienist. They stimulate discussion concerning the inter- pertive and cognitive skill required of health professional (10:18) Orig. air date: JAN 17 75Comstock, Frank; Chasteen, Joseph, "Clean Techniques": A series of five clean technique demonstrations: (16:12) Orig. air date: SEP 10 75White, Stewart L., "Computers In The Pilot Program in Dentistry": Discusses the current and future uses of the computer, cost effectveness, and the potential for sharing. (25:45) Orig. air date: MAR 9 77Eddlemon, Charles; German, Marjorie, "Introduction to the Pilot Program in Dentistry": Discussion of progression of Pilot Program in Dentistry from its inception to present. (4:05) Orig. air date: MAY 11 76Eddlemon, Chuck; Mevis, Howard, "Achievements in the Pilot Program in Dentistry": A program describing methods in instructional design and development.Two courses that were developed at the University of Michigan School of Dentistry are presented . (19:25) Orig. air date: FEB 13 75Miller, Doris, "Trigger Tapes - Legal & Ethical Questions": Presents several vignettes that can be used to trigger"""" discussions in Dental health care professionals concerning legal and ethical questions in the Dental practice. (7:06) Orig. air date: MAR 6 75http://deepblue.lib.umich.edu/bitstream/2027.42/64983/2/102-3.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/3/102-5.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/4/102-6.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/5/134-3-4UnderstandSkills.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/6/136-3TriggerTapes.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/7/138-1TriggerTapes.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/8/177TriggerTapes.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/9/182-3Inconsistency.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/10/225-1DentalUnit.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/11/225-2PatientEducation.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/12/226-2PatienEducation.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/13/261-2OralHygiene.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/14/319-3PatientEducation.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/15/338-1CleanTechniques.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/16/339-2Computers.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/17/376-3Intro2PPD.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/18/411-2PPD.movhttp://deepblue.lib.umich.edu/bitstream/2027.42/64983/19/415-2TriggerTapes.mo