bluuebunny/crossref_metadata_2025_split huggingface dataset:
Exploring protein interfaces with a general photochemical reagent
Authors: Gómez Gabriela E. , Cauerhff Ana , Craig Patricio O. , Goldbaum Fernando A. , Delfino José M. 
Published: April 2006

AbstractProtein folding, natural conformational changes, or interaction between partners involved in recognition phenomena brings about differences in the solvent‐accessible surface area (SASA) of the polypeptide chain. This primary event can be monitored by the differential chemical reactivity of functional groups along the protein sequence. Diazirine (DZN), a photoreactive gas similar in size to water, generates methylene carbene (:CH2). The extreme chemical reactivity of this species allows the almost instantaneous and indiscriminate modification of its immediate molecular cage. 3H‐DZN was successfully used in our laboratory for studying protein structure and folding. Here we address for the first time the usefulness of this probe to examine the area of interaction in protein–protein complexes. For this purpose we chose the complex formed between hen egg white lysozyme (HEWL) and the monoclonal antibody IgG1 D1.3. :CH2 labeling of free HEWL or complexed with IgG1 D1.3 yields 2.76 and 2.32 mmol CH2 per mole protein at 1 mM DZN concentration, respectively. This reduction (15%) becomes consistent with the expected decrement in the SASA of HEWL occurring upon complexation derived from crystallographic data (11%), in agreement with the known unspecific surface labeling reaction of :CH2. Further comparative analysis at the level of tryptic peptides led to the identification of the sites involved in the interaction. Remarkably, those peptides implicated in the contact area show the highest differential labeling: H15GLDNYR21, G117TDVQAWIR125, andG22YSLGNWVCAAK33. Thus, protein footprinting with DZN emerges as a feasible methodology useful for mapping contact regions of protein domains involved in macromolecular assemblies.
Solution structure of Asl1650, an acyl carrier protein from Anabaena sp. PCC 7120 with a variant phosphopantetheinylation‐site sequence
Authors: Johnson Margaret A. , Peti Wolfgang , Herrmann Torsten , Wilson Ian A. , Wüthrich Kurt 
Published: April 2006

AbstractCyanobacteria, such as Anabaena, produce a variety of bioactive natural products via polyketide synthases (PKS), nonribosomal peptide synthetases (NRPS), and hybrid peptide/polyketide pathways. The protein Asl1650, which is a member of the acyl carrier protein family from the cyanobacterium Anabaena sp. PCC 7120, is encoded in a region of the Anabaena genome that is rich in PKS and NRPS genes. To gain new insight into the physiological role of acyl carriers in Anabaena, the solution structure of Asl1650 has been solved by NMR spectroscopy. The protein adopts a twisted antiparallel four‐helix bundle fold, with a variant phosphopantetheine‐attachment motif positioned at the start of the second helix. Structure comparisons with proteins from other organisms suggest a likely physiological function as a discrete peptidyl carrier protein.
Rapid peptide‐based screening on the substrate specificity of severe acute respiratory syndrome (SARS) coronavirus 3C‐like protease by matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry
Authors: Chu Ling‐Hon Matthew , Choy Wai‐Yan , Tsai Sau‐Na , Rao Zihe , Ngai Sai‐Ming 
Published: April 2006

AbstractSevere acute respiratory syndrome coronavirus (SARS‐CoV) 3C‐like protease (3CLpro) mediates extensive proteolytic processing of replicase polyproteins, and is considered a promising target for anti‐SARS drug development. Here we present a rapid and high‐throughput screening method to study the substrate specificity of SARS‐CoV 3CLpro. Six target amino acid positions flanking the SARS‐CoV 3CLpro cleavage site were investigated. Each batch of mixed peptide substrates with defined amino acid substitutions at the target amino acid position was synthesized via the “cartridge replacement” approach and was subjected to enzymatic cleavage by recombinant SARS‐CoV 3CLpro. Susceptibility of each peptide substrate to SARS‐CoV 3CLpro cleavage was monitored simultaneously by matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometry (MALDI‐TOF MS). The hydrophobic pocket in the P2 position at the protease cleavage site is crucial to SARS‐CoV 3CLpro‐specific binding, which is limited to substitution by hydrophobic residue. The binding interface of SARS‐CoV 3CLpro that is facing the P1′ position is suggested to be occupied by acidic amino acids, thus the P1′ position is intolerant to acidic residue substitution, owing to electrostatic repulsion. Steric hindrance caused by some bulky or β‐branching amino acids in P3 and P2′ positions may also hinder the binding of SARS‐CoV 3CLpro. This study generates a comprehensive overview of SARS‐CoV 3CLpro substrate specificity, which serves as the design basis of synthetic peptide‐based SARS‐CoV 3CLpro inhibitors. Our experimental approach is believed to be widely applicable for investigating the substrate specificity of other proteases in a rapid and high‐throughput manner that is compatible for future automated analysis.
Affinity enhancement of an in vivo matured therapeutic antibody using structure‐based computational design
Authors: Clark Louis A. , Boriack‐Sjodin P. Ann , Eldredge John , Fitch Christopher , Friedman Bethany , Hanf Karl J.M. , Jarpe Matthew , Liparoto Stefano F. , Li You , Lugovskoy Alexey , Miller Stephan , Rushe Mia , Sherman Woody , Simon Kenneth , Van Vlijmen Herman 
Published: April 2006

AbstractImproving the affinity of a high‐affinity protein–protein interaction is a challenging problem that has practical applications in the development of therapeutic biomolecules. We used a combination of structure‐based computational methods to optimize the binding affinity of an antibody fragment to the I‐domain of the integrin VLA1. Despite the already high affinity of the antibody (Kd ∼7 nM) and the moderate resolution (2.8 Å) of the starting crystal structure, the affinity was increased by an order of magnitude primarily through a decrease in the dissociation rate. We determined the crystal structure of a high‐affinity quadruple mutant complex at 2.2 Å. The structure shows that the design makes the predicted contacts. Structural evidence and mutagenesis experiments that probe a hydrogen bond network illustrate the importance of satisfying hydrogen bonding requirements while seeking higher‐affinity mutations. The large and diverse set of interface mutations allowed refinement of the mutant binding affinity prediction protocol and improvement of the single‐mutant success rate. Our results indicate that structure‐based computational design can be successfully applied to further improve the binding of high‐affinity antibodies.
Thermodynamics of unfolding of an integral membrane protein in mixed micelles
Authors: Sehgal Pankaj , Otzen Daniel E. 
Published: April 2006

AbstractQuantitative studies of membrane protein folding and unfolding can be difficult because of difficulties with efficient refolding as well as a pronounced propensity to aggregate. However, mixed micelles, consisting of the anionic detergent sodium dodecyl sulfate and the nonionic detergent dodecyl malto‐side facilitate reversible and quantitative unfolding and refolding. The 4‐transmembrane helix protein DsbB from the inner membrane of Escherichia coli unfolds in mixed micelles according to a three‐state mechanism involving an unfolding intermediate I. The temperature dependence of the kinetics of this reaction between 15° and 45°C supports that unfolding from I to the denatured state D is accompanied by a significant decrease in heat capacity. For water‐soluble proteins, the heat capacity increases upon unfolding, and this is generally interpreted as the increased binding of water to the protein as it unfolds, exposing more surface area. The decrease in DsbB's heat capacity upon unfolding is confirmed by independent thermal scans. The decrease in heat capacity is not an artifact of the use of mixed micelles, since the water soluble protein S6 shows conventional heat‐capacity changes in detergent. We speculate that it reflects the binding of SDS to parts of DsbB that are solvent‐exposed in the native DM‐bound state. This implies that the periplasmic loops of DsbB are relatively unstructured. This anomalous thermodynamic behavior has not been observed for β‐barrel membrane proteins, probably because they do not bind SDS so extensively. Thus the thermodynamic behavior of membrane proteins appears to be intimately connected to their detergent‐binding properties.
The contribution of the residues from the main hydrophobic core of ribonuclease A to its pressure‐folding transition state
Authors: Font Josep , Benito Antoni , Lange Reinhard , Ribó Marc , Vilanova Maria 
Published: April 2006

AbstractThe role of hydrophobic interactions established by the residues that belong to the main hydrophobic core of ribonuclease A in its pressure‐folding transition state was investigated using the Φ‐value method. The folding kinetics was studied using pressure‐jump techniques both in the pressurization and depressurization directions. The ratio between the folding activation volume and the reaction volume (βp‐value), which is an index of the compactness or degree of solvation of the transition state, was calculated. All the positions analyzed presented fractional Φf‐values, and the lowest were those corresponding to the most critical positions for the ribonuclease A stability. The structure of the transition state of the hydrophobic core of ribonuclease A, from the point of view of formed interactions, is a relatively, uniformly expanded form of the folded structure with a mean Φf‐value of 0.43. This places it halfway between the folded and unfolded states. On the other hand, for the variants, the average of βp‐values is 0.4, suggesting a transition state that is 40% native‐like. Altogether the results suggest that the pressure‐folding transition state of ribonuclease A looks like a collapsed globule with some secondary structure and a weakened hydrophobic core. A good correlation was found between the Φf‐values and the Δβp‐values. Although the nature of the transition state inferred from pressure‐induced folding studies and the results of the protein engineering method have been reported to be consistent for other proteins, to the best of our knowledge this is the first direct comparison using a set of mutants.
The C‐terminal domain of the transcriptional corepressor CtBP is intrinsically unstructured
Authors: Nardini Marco , Svergun Dmitri , Konarev Peter V. , Spanò Stefania , Fasano Mauro , Bracco Chiara , Pesce Alessandra , Donadini Alessandra , Cericola Claudia , Secundo Francesco , Luini Alberto , Corda Daniela , Bolognesi Martino 
Published: April 2006

AbstractC‐terminal binding proteins (CtBPs) are moonlighting proteins involved in nuclear transcriptional corepression and in Golgi membrane tubule fission. Structural information on CtBPs is available for their substrate‐binding domain, responsible for transcriptional repressor recognition/binding, and for the nucleotide‐binding domain, involved in NAD(H)‐binding and dimerization. On the contrary, little is known about the structure of CtBP C‐terminal region (∼90 residues), hosting sites for post‐translational modifications. In the present communication we apply a combined approach based on bioinformatics, nuclear magnetic resonance, circular dichroism spectroscopy, and small‐angle X‐ray scattering, and we show that the CtBP C‐terminal region is intrinsically unstructured in the full‐length CtBP and in constructs lacking the substrate‐ and/or the nucleotide‐binding domains. The flexible nature of this protein region, and its structural transitions, may be instrumental for CtBP recognition and binding to diverse molecular partners.
ULTRASTRUCTURAL MORPHOLOGY OF MICROTHAMNION ZOOSPORES1
Authors: Watson Melvin William , Arnott Howard J. 
Published: April 2006

SUMMARYThe Ultrastructure of Microthamnion zoospores is described (exclusive of the finer details of the flagellar apparatus). The zoospores have a typical chlorophycean morphology but, in addition, many unique features. The chloroplasts contain starch but no pyrenoid. Thylakoids may run from one edge of the chloroplast to the other and usually anastomose into 2‐ to 8‐membered stacks. The internal morphology is highly polarized and characterized by an intimate proximity and constant spacing between many organelle membranes. All the organelles are asymmetrically distributed within the cell in a precise manner. The anterior region of the zoospore is attenuated into a neck which contains a single, massive mitochondrion. A fibrous rhizoplast lies beside the mitochondrion and appears to connect the flagellar apparatus directly with the outer membrane of the nuclear envelope. In addition, this outer membrane is extended over a distance of several microns to eventually lie in close proximity to the basal bodies. Oil vacuoles and lipid bodies are restricted to the posterior end of the zoospore.
ULTRASTRUCTURE OF TETRASPOROGENESIS IN THE MARINE RED ALGA PTILOTA HYPNOIDES1
Authors: Scott J. L. , Dixon P. S. 
Published: April 2006

SUMMARYBoth tetrasporangia and dormant apical cells of short vegetative filaments of the marine red alga Ptilota hypnoides have been examined by electron microscopy. Various cytoplasmic inclusions readily distinguish the vegetative apical cells from the reproductive apical cells which become tetrasporangial mother cells. The transformation of tetrasporangial mother cells into mature tetrasporangia involves a series of cytoplasmic changes which can be correlated with specific changes in the investing wall layers. The extracellular changes provide the basic criteria for the division of tetrasporogenesis into 3 successive stages. The ultrastructure of each stage is described and discussed in relation to the current knowledge of red algal cytology. In addition, a possible mechanism for the liberation of spores and gametes of red algae is proposed.
GROWTH OF THE GREEN ALGA CODIUM FRAGILE IN A CONNECTICUT ESTUARY1
Authors: Malinowski K. C. , Ramus J. 
Published: April 2006

SUMMARYAn in situ comparison of environmental and physiological factors was undertaken in 1971–72 (15 months) in the Niantic River estuary to elucidate some of the important aspects, of the growth and development of the seaweed Codium fragile.In general, Codium in the estuary has a growing season of from 6 to 9 months. Growth increments during this period are relatively constant. Temperature and salinity are the main limiting factors to growth, although low summer concentrations of inorganic nitrogen may also be involved. Reproduction by means of swarmers occurs only in late summer or autumn. Codium appears highly adapted to the role of a colonizing species. It not only possesses the capability to occupy the harshest of environments, but also a system of reproductive alternatives which facilitates rapid colonization. The availability of substrate for attachment is the chief factor limiting its spread in this estuarine system.
Nurse‐related factors in the delivery of preoperative patient education
Authors: Fitzpatrick Elizabeth , Hyde Abbey 
Published: April 2006

Aims and objectives.  This article aimed to explore the factors relating to nurses themselves that influence the delivery of preoperative patient education in everyday surgical clinical contexts at one large general adult teaching hospital in Ireland.Background.  As landmark studies of preoperative education undertaken in the UK in the 1970s identified the superiority of structured programmes of patient education over and above ‘regular’ preoperative care, there have been many intervention/outcome experimental studies carried out in this area. However, there has been little interpretative work conducted that explores the regular, or ‘usual’ preoperative education given to patients in everyday surgical units.Methods.  A sample of 12 experienced surgical nurses was selected and each participant was interviewed in depth. Data were analysed using a qualitative strategy resembling grounded theory.Results.  Findings indicate that preoperative education was variously interpreted by participants, and participants’ accounts suggested that different understandings and practices by nurses resulted in patients receiving different levels of care. In addition, diverse levels of knowledge and experience of individual nurses resulted in unevenness in the type of preoperative education that patients received. A number of participants advocated a more formal method of preparation for nurses in the area of preoperative education. Finally, in some surgical areas, specialist nurses worked side‐by‐side with regular ward nurses and their input in preoperative education was largely seen by participants in a positive way, particularly in view of the structural constraints that ward nurses faced.Conclusions.  We conclude that nurse‐related factors in preoperative education do not arise in a vaccum, but rather are related to the wider organizational practices and culture. The difficulties with preoperative education identified in data may be addressed through organizational investment in preoperative education.Relevance to clinical practice.  This paper produces evidence for nurses and clinical nurse managers about how nurse‐related factors impede preoperative education, and may provide a starting point for how to begin to address obstacles to better preoperative care.
Supplementary ultraviolet‐B irradiation reveals differences in stress responses between Arabidopsis thaliana ecotypes*
Authors: KALBINA IRINA , STRID ÅKE 
Published: April 2006

ABSTRACTIrradiation of Arabidopsis thaliana ecotypes C24, Wassilewskija (Ws) and Columbia‐0 (Col‐0) with supplementary ultraviolet‐A+B (UV‐A+B) radiation revealed ecotype‐specific differences in expression of the gene for the pathogenesis‐related protein PR‐5. C24 showed an increased expression level of PR‐5 (5‐ and 20‐fold higher compared with Col‐0 and Ws, respectively). Expression of other molecular markers such as CHS (encoding chalcone synthase), MEB5.2[encoding a gene strongly up‐regulated by ultraviolet‐B (UV‐B)] and PYROA[encoding a pyridoxine (Vitamin B6) biosynthesis enzyme] only showed slight differences between ecotypes. Oxidative stress during UV‐A+B exposure was monitored by staining for H2O2. This analysis also revealed important ecotype‐specific differences. ‘H2O2 hot spots’ were found in C24, whereas an even distribution of H2O2 was found in Ws and Col‐0. Necrotic lesions also appeared on C24 leaves after prolonged UV‐B exposure. There was a reverse correlation between the H2O2 steady‐state concentration and the PR‐5 gene expression; Ws showed the highest level of H2O2 accumulation but the lowest expression level of the PR‐5 gene. Furthermore, application of paraquat on the rosettes led to similar PR‐5 expression and H2O2 accumulation patterns as were found after UV‐A+B irradiation. The observed ecotypic differences were also reflected in a statistically significant UV‐B‐dependent decrease in biomass, rosette size and leaf area for Ws, but not for C24 and Col‐0. Our results show that a significant ecotype‐specific genetic variability in general UV‐B responses in Arabidopsis exists. Moreover, the signal transduction or gene regulation pathway for PR‐5 differs from the other molecular markers used in this study.
Specialist Training at the Faculty of Odontology, Malmö University
Authors: Lindh C , Lindberg P , Rohlin M. 
Published: April 2006

Aim  To describe the specialist training at the Faculty of Odontology in Malmö for eight specialities that are certified in Sweden and report the results of a questionnaire among former specialist trainees.Material and Method  The objectives of the specialist training are set on three levels: the national level according to the Swedish National Board for Health and Welfare, the faculty level, the so called Malmö model, and finally the specialist subject level. The specialist training in Malmö is implemented in collaboration with the Public Dental Health Care in southern Sweden. The Postgraduate Education Board of the faculty scrutinizes and approves each trainee's individualized specialist training programme. The clinical part of the training is to be shared between the collaborating units and the theoretical part is the prime responsibility of the faculty. During the three‐year‐programme, seven core courses are held for all trainees irrespective of specialist subject parallel to the clinical training. Every semester there are two joint seminars: one seminar on a current odontological theme and one on the following subjects; epidemiology, ethics, migration and ethnic relations, environment, gender, leadership. A written and oral programme evaluation is performed yearly by the Chair of the Postgraduate Education Board together with a representative of the Public Dental Health Care. As part of the 2003 evaluation a questionnaire was sent to 31 specialist graduates trained between 2000 and 2003. Twenty‐four specialists responded, 13 graduates from Malmö and 11 from other Swedish specialist institutions.Results and Conclusion  A majority of the 24 responding graduates express that the work as specialists suit them very well and that the education well serve the purpose to work as a specialist. A majority of the graduates from Malmö were positive to the joint courses during their training. Some thought that there should be more joint activities among trainees of different subject specialities.
Socio‐economic Factors Guiding the Number of Dental Specialists in Europe
Authors: Widstrom E 
Published: April 2006

In the European Economic area, four countries have no recognised dental specialties and eight have five or more. Orthodontics is recognised in most countries and Oral Maxillofacial Surgery and/or Oral Surgery in over 60%. Other common specialties are Paedodontics, Periodontics and Prosthetics. Variation in the numbers of specialties and specialists between individual countries can partly be explained by oral healthcare systems. A further factor is that the population has become older, wealthier, less edentulous and demands more advanced treatment. Those who support an increase in dental specialties and in numbers of specialists, highlight professional development and better care for patients. Those against are afraid of restrictions in general practice and competition.Most existing specialties seem to have emerged from professional interests. Only two, Orthodontics and Oral Surgery, have been formally recognised by the European Commission (EC).A new EC Draft Directive proposes that specialties, which exist in 10 or more Member States will be recognised by the EU. This could lead to more dental specialties. The need for specialists should be assessed on the basis of treatment needs of the populations concerned. Key questions are how to ensure that, at present and in the future, specialties and specialists serve these needs and offer high quality and affordable services.