Texture perception determined by doy protein isolate and inulin addition in potato puree: links with mechanical and microstructural features

This study evaluated the effect of adding soy protein isolate (SPI) and long-chain perception, trained and untrained panel inulin (INL) blends with 10 different SPI : INL ratios on the textural, rheological and 17 microstructural properties of freshly made and frozen/thawed potato puree. All the potato puree samples were subjected to a sensory texture pro?le analysis and a 21 trained panel rated the intensity of six descriptors, while an untrained panel did the same on six selected frozen/thawed products. The main SPI : INL ratio effect remained signi?cant for all the descriptors evaluated, when the analysis of variance was applied considering the untrained assessors as random effects. However, only trained panel scores for creaminess corresponded well with untrained assessor. Rheological ?ow index values were linked with variations in perceived consistency, and geometric and surface textural attributes were explained by structural features such as the presence of INL crystallites and SPI coarse strands.

Microstructural segmentation of 3D images of alsphalt specimen using Matlab engine

En este proyecto se ha desarrollado un código de MATLAB para el procesamiento de imágenes tomográficas 3D, de muestras de asfalto de carreteras en Polonia. Estas imágenes en 3D han sido tomadas por un equipo de investigación de la Universidad Tecnológica de Lodz (LUT). El objetivo de este proyecto es crear una herramienta que se pueda utilizar para estudiar las diferentes muestras de asfalto 3D y pueda servir para estudiar las pruebas de estrés que experimentan las muestras en el laboratorio. Con el objetivo final de encontrar soluciones a la degradación sufrida en las carreteras de Polonia, debido a diferentes causas, como son las condiciones meteorológicas. La degradación de las carreteras es un tema que se ha investigado desde hace muchos años, debido a la fuerte degradación causada por diferentes factores como son climáticos, la falta de mantenimiento o el tráfico excesivo en algunos casos. Es en Polonia, donde estos tres factores hacen que la composición de muchas carreteras se degrade rápidamente, sobre todo debido a las condiciones meteorológicas sufridas a lo largo del año, con temperaturas que van desde 30° C en verano a -20° C en invierno. Esto hace que la composición de las carreteras sufra mucho y el asfalto se levante, lo que aumenta los costos de mantenimiento y los accidentes de carretera. Este proyecto parte de la base de investigación que se lleva a cabo en la LUT, tratando de mejorar el análisis de las muestras de asfalto, por lo que se realizarán las pruebas de estrés y encontrar soluciones para mejorar el asfalto en las carreteras polacas. Esto disminuiría notablemente el costo de mantenimiento. A pesar de no entrar en aspectos muy técnicos sobre el asfalto y su composición, se ha necesitado realizar un estudio profundo sobre todas sus características, para crear un código capaz de obtener los mejores resultados. Por estas razones, se ha desarrollado en Matlab, los algoritmos que permiten el estudio de los especímenes 3D de asfalto. Se ha utilizado este software, ya que Matlab es una poderosa herramienta matemática que permite operar con matrices para realización de operaciones rápidamente, permitiendo desarrollar un código específico para el tratamiento y procesamiento de imágenes en 3D. Gracias a esta herramienta, estos algoritmos realizan procesos tales como, la segmentación de la imagen 3D, pre y post procesamiento de la imagen, filtrado o todo tipo de análisis microestructural de las muestras de asfalto que se están estudiando. El código presentado para la segmentación de las muestras de asfalto 3D es menos complejo en su diseño y desarrollo, debido a las herramientas de procesamiento de imágenes que incluye Matlab, que facilitan significativamente la tarea de programación, así como el método de segmentación utilizado. Respecto al código, este ha sido diseñado teniendo en cuenta el objetivo de facilitar el trabajo de análisis y estudio de las imágenes en 3D de las muestras de asfalto. Por lo tanto, el principal objetivo es el de crear una herramienta para el estudio de este código, por ello fue desarrollado para que pueda ser integrado en un entorno visual, de manera que sea más fácil y simple su utilización. Ese es el motivo por el cual todos estos algoritmos y funciones, que ha sido desarrolladas, se integrarán en una herramienta visual que se ha desarrollado con el GUIDE de Matlab. Esta herramienta ha sido creada en colaboración con Jorge Vega, y fue desarrollada en su proyecto final de carrera, cuyo título es: Segmentación microestructural de Imágenes en 3D de la muestra de asfalto utilizando Matlab. En esta herramienta se ha utilizado todo las funciones programadas en este proyecto, y tiene el objetivo de desarrollar una herramienta que permita crear un entorno gráfico intuitivo y de fácil uso para el estudio de las muestras de 3D de asfalto. Este proyecto se ha dividido en 4 capítulos, en un primer lugar estará la introducción, donde se presentarán los aspectos más importante que se va a componer el proyecto. En el segundo capítulo se presentarán todos los datos técnicos que se han tenido que estudiar para desarrollar la herramienta, entre los que cabe los tres temas más importantes que se han estudiado en este proyecto: materiales asfálticos, los principios de la tomografías 3D y el procesamiento de imágenes. Esta será la base para el tercer capítulo, que expondrá la metodología utilizada en la elaboración del código, con la explicación del entorno de trabajo utilizado en Matlab y todas las funciones de procesamiento de imágenes utilizadas. Además, se muestra todo el código desarrollado, así como una descripción teórica de los métodos utilizados para el pre-procesamiento y segmentación de las imagenes en 3D. En el capítulo 4, se mostrarán los resultados obtenidos en el estudio de una de las muestras de asfalto, y, finalmente, el último capítulo se basa en las conclusiones sobre el desarrollo de este proyecto. En este proyecto se ha llevado han realizado todos los puntos que se establecieron como punto de partida en el anteproyecto para crear la herramienta, a pesar de que se ha dejado para futuros proyectos nuevas posibilidades de este codigo, como por ejemplo, la detección automática de las diferentes regiones de una muestra de asfalto debido a su composición. Como se muestra en este proyecto, las técnicas de procesamiento de imágenes se utilizan cada vez más en multitud áreas, como pueden ser industriales o médicas. En consecuencia, este tipo de proyecto tiene multitud de posibilidades, y pudiendo ser la base para muchas nuevas aplicaciones que se puedan desarrollar en un futuro. Por último, se concluye que este proyecto ha contribuido a fortalecer las habilidades de programación, ampliando el conocimiento de Matlab y de la teoría de procesamiento de imágenes. Del mismo modo, este trabajo proporciona una base para el desarrollo de un proyecto más amplio cuyo alcance será una herramienta que puedas ser utilizada por el equipo de investigación de la Universidad Tecnológica de Lodz y en futuros proyectos. ABSTRACT In this project has been developed one code in MATLAB to process X-ray tomographic 3D images of asphalt specimens. These images 3D has been taken by a research team of the Lodz University of Technology (LUT). The aim of this project is to create a tool that can be used to study differents asphalt specimen and can be used to study them after stress tests undergoing the samples. With the final goal to find solutions to the degradation suffered roads in Poland due to differents causes, like weather conditions. The degradation of the roads is an issue that has been investigated many years ago, due to strong degradation suffered caused by various factors such as climate, poor maintenance or excessive traffic in some cases. It is in Poland where these three factors make the composition of many roads degrade rapidly, especially due to the weather conditions suffered along the year, with temperatures ranging from 30 o C in summer to -20 ° C in winter. This causes the roads suffers a lot and asphalt rises shortly after putting, increasing maintenance costs and road accident. This project part of the base that research is taking place at the LUT, in order to better analyze the asphalt specimens, they are tested for stress and find solutions to improve the asphalt on Polish roads. This would decrease remarkable maintenance cost. Although this project will not go into the technical aspect as asphalt and composition, but it has been required a deep study about all of its features, to create a code able to obtain the best results. For these reasons, there have been developed in Matlab, algorithms that allow the study of 3D specimens of asphalt. Matlab is a powerful mathematical tool, which allows arrays operate fastly, allowing to develop specific code for the treatment and processing of 3D images. Thus, these algorithms perform processes such as the multidimensional matrix sgementation, pre and post processing with the same filtering algorithms or microstructural analysis of asphalt specimen which being studied. All these algorithms and function that has been developed to be integrated into a visual tool which it be developed with the GUIDE of Matlab. This tool has been created in the project of Jorge Vega which name is: Microstructural segmentation of 3D images of asphalt specimen using Matlab engine. In this tool it has been used all the functions programmed in this project, and it has the aim to develop an easy and intuitive graphical environment for the study of 3D samples of asphalt. This project has been divided into 4 chapters plus the introduction, the second chapter introduces the state-of-the-art of the three of the most important topics that have been studied in this project: asphalt materials, principle of X-ray tomography and image processing. This will be the base for the third chapter, which will outline the methodology used in developing the code, explaining the working environment of Matlab and all the functions of processing images used. In addition, it will be shown all the developed code created, as well as a theoretical description of the methods used for preprocessing and 3D image segmentation. In Chapter 4 is shown the results obtained from the study of one of the specimens of asphalt, and finally the last chapter draws the conclusions regarding the development of this project.

PREDIRCAM eHealth platform for individualized telemedical assistance for lifestyle modification in the treatment of obesity, diabetes, and cardiometabolic risk prevention: a pilot study (PREDIRCAM 1)

Background: Healthy diet and regular physical activity are powerful tools in reducing diabetes and cardiometabolic risk. Various international scientific and health organizations have advocated the use of new technologies to solve these problems. The PREDIRCAM project explores the contribution that a technological system could offer for the continuous monitoring of lifestyle habits and individualized treatment of obesity as well as cardiometabolic risk prevention. Methods: PREDIRCAM is a technological platform for patients and professionals designed to improve the effectiveness of lifestyle behavior modifications through the intensive use of the latest information and communication technologies. The platform consists of a web-based application providing communication interface with monitoring devices of physiological variables, application for monitoring dietary intake, ad hoc electronic medical records, different communication channels, and an intelligent notification system. A 2-week feasibility study was conducted in 15 volunteers to assess the viability of the platform. Results: The website received 244 visits (average time/session: 17 min 45 s). A total of 435 dietary intakes were recorded (average time for each intake registration, 4 min 42 s ± 2 min 30 s), 59 exercises were recorded in 20 heart rate monitor downloads, 43 topics were discussed through a forum, and 11 of the 15 volunteers expressed a favorable opinion toward the platform. Food intake recording was reported as the most laborious task. Ten of the volunteers considered long-term use of the platform to be feasible. Conclusions: The PREDIRCAM platform is technically ready for clinical evaluation. Training is required to use the platform and, in particular, for registration of dietary food intake.

Influencia de la porosidad y su fase acuosa en la estabilidad microstructural de la pasta de cemento, mortero y hormigón

Se estudia la importancia de la microestructura: porosidad y fase acuosa contenida en los poros en relación con las propiedades de la pasta de cemento, mortero y hormigón, así como las consecuencias de sus alteraciones en la estabilidad del material.

Microstructural and mechanical characteristics of W-2Ti and W-1TiC processed by Hot Isostatic Pressing

It has been demonstrated that mechanical alloying and subsequent consolidation by hot isostatic pressing (HIP) is a successful route to produce dispersion strengthened W alloys with properties satisfying the design requirements of particular plasma facing components in the fusion reactor. However, the presence of the alloying element as a phase filling large interstices between W particles appears to reduce the mechanical properties of these alloys. In order to limit this phase separation induced by the HIP treatment and the detrimental effects on the mechanical properties, the enhancement of the mechanical alloying process, and the effect of a postconsolidation heat treatment in an reducing atmosphere, have been investigated.

Microstructural and mechanical characterization of annealed tungsten (W) and potassium-doped tungsten foils

Here we show that potassium-doped tungsten foil should be preferred to pure tungsten foil when considering tungsten laminate pipes for structural divertor applications. Potassium-doped tungsten materials are well known from the bulb industry and show an enhanced creep and recrystallization behaviour that can be explained by the formation of potassium-filled bubbles that are surrounding the elongated grains, leading to an interlocking of the microstructure. In this way, the ultra-fine grained (UFG) microstructure of tungsten foil can be stabilized and with it the extraordinary mechanical properties of the foil in terms of ductility, toughness, brittle-to-ductile transition, and radiation resistance. In this paper we show the results of three-point bending tests performed at room temperature on annealed pure tungsten and potassium-doped tungsten foils (800, 900, 1000, 1100, 1200, 1300, 1400, 1600, 1800, 2000, 2200, and 2400 °C for 1 h in vacuum). The microstructural assessment covers the measurement of the hardness and analyses of fractured surfaces as well as a comparison of the microstructure by optical microscopy. The results show that there is a positive effect of potassium-doped tungsten foils compared to pure tungsten foil and demonstrate the potential of the doped foil

Microstructural and mechanical characteristics of W-2Ti and W-1TiC processed by hot isostatic pressing

W–2Ti and W–1TiC alloys were produced by mechanical alloying and consolidation by hot isostatic pressing. The composition and microstructural characteristics of these alloys were studied by X-ray diffraction, energy dispersion spectroscopy and scanning electron microscopy. The mechanical behavior of the consolidated alloys was characterized by microhardness measurements and three point bending tests. The mechanical characteristics of the W–2Ti alloy appear to be related to solution hardening. In W–1TiC, the residual porosity should be responsible for the poor behavior observed in comparison with W–2Ti.

Thermoelectric assessment of laser peening induced effects on a metallic biomedical Ti6Al4V

Laser peening has recently emerged as a useful technique to overcome detrimental effects associated to another well-known surface modification processes such as shot peening or grit blasting used in the biomedical field. It is worth to notice that besides the primary residual stress effect, thermally induced effects might also cause subtle surface and subsurface microstructural changes that might influence corrosion resistance. Moreover, since maximum loads use to occur at the surface, they could also play a critical role in the fatigue strength. In this work, plates of Ti-6Al-4V alloy of 7 mm in thickness were modified by laser peening without using a sacrificial outer layer. Irradiation by a Q-switched Nd-YAG laser (9.4 ns pulse length) working in fundamental harmonic at 2.8 J/pulse and with water as confining medium was used. Laser pulses with a 1.5 mm diameter at an equivalent overlapping density (EOD) of 5000 cm-2 were applied. Attempts to analyze the global induced effects after laser peening were addressed by using the contacting and non-contacting thermoelectric power (TEP) techniques. It was demonstrated that the thermoelectric method is entirely insensitive to surface topography while it is uniquely sensitive to subtle variations in thermoelectric properties, which are associated with the different material effects induced by different surface modification treatments. These results indicate that the stress-dependence of the thermoelectric power in metals produces sufficient contrast to detect and quantitatively characterize regions under compressive residual stress based on their thermoelectric power contrast with respect to the surrounding intact material. However, further research is needed to better separate residual stress effects from secondary material effects, especially in the case of low-conductivity engineering materials like titanium alloys.

Microstructural and mechanical characterization of tungsten based materials for fusion reactors

El wolframio (W) y sus aleaciones se consideran los mejores candidatos para la construcción del divertor en la nueva generación de reactores de fusión nuclear. Este componente va a recibir las cargas térmicas más elevadas durante el funcionamiento del reactor ya que estará en contacto directo con el plasma. En los últimos años, después de un profundo análisis y siguiendo una estrategia de reducción de costes, la Organización de ITER tomó la decisión de construir el divertor integramente de wolframio desde el principio. Por ello, el wolframio no sólo actuará como material en contacto con el plasma (PFM), sino que también tendría aplicaciones estructurales. El wolframio, debido a sus excelentes propiedades termo-físicas, cumple todos los requerimientos para ser utilizado como PFM, sin embargo, su inherente fragilidad pone en peligro su uso estructural. Por tanto, uno de los principales objetivos de esta tesis es encontrar una aleación de wolframio con menor fragilidad. Durante éste trabajo, se realizó la caracterización microstructural y mecánica de diferentes materiales basados en wolframio. Sin embargo, ésta tarea es un reto debido a la pequeña cantidad de material suministrado, su reducido tamaño de grano y fragilidad. Por ello, para una correcta medida de todas las propiedades físicas y mecánicas se utilizaron diversas técnicas experimentales. Algunas de ellas se emplean habitualmente como la nanoindentación o los ensayos de flexión en tres puntos (TPB). Sin embargo, otras fueron especificamente desarrolladas e implementadas durante el desarrollo de esta tesis como es el caso de la medida real de la tenacidad de fractura en los materiales masivos, o de las medidas in situ de la tenacidad de fractura en las láminas delgadas de wolframio. Diversas composiciones de aleaciones de wolframio masivas (W-1% Y2O3, W-2% V-0.5% Y2O3, W-4% V-0.5% Y2O3, W-2% Ti-1% La2O3 y W-4% Ti-1% La2O3) se han estudiado y comparado con un wolframio puro producido en las mismas condiciones. Estas aleaciones, producidas por ruta pulvimetalúrgica de aleado mecánico (MA) y compactación isostática en caliente (HIP), fueron microstructural y mecánicamente caracterizadas desde 77 hasta 1473 K en aire y en alto vacío. Entre otras propiedades físicas y mecánicas se midieron la dureza, el módulo elástico, la resistencia a flexión y la tenacidad de fractura para todas las aleaciones. Finalmente se analizaron las superficies de fractura después de los ensayos de TPB para relacionar los micromecanismos de fallo con el comportamiento macroscópico a rotura. Los resultados obtenidos mostraron un comportamiento mecánico frágil en casi todo el intervalo de temperaturas y para casi todas las aleaciones sin mejoría de la temperatura de transición dúctil-frágil (DBTT). Con el fin de encontrar un material base wolframio con una DBTT más baja se realizó también un estudio, aún preliminar, de láminas delgadas de wolframio puro y wolframio dopado con 0.005wt.% potasio (K). Éstas láminas fueron fabricadas industrialmente mediante sinterizado y laminación en caliente y en frío y se sometieron posteriormente a un tratamiento térmico de recocido desde 1073 hasta 2673 K. Se ha analizado la evolución de su microestructura y las propiedades mecánicas al aumentar la temperatura de recocido. Los resultados mostraron la estabilización de los granos de wolframio con el incremento de la temperatura de recocido en las láminas delgadas de wolframio dopado con potasio. Sin embargo, es necesario realizar estudios adicionales para entender mejor la microstructura y algunas propiedades mecánicas de estos materiales, como la tenacidad de fractura. Tungsten (W) and tungsten-based alloys are considered to be the best candidate materials for fabricating the divertor in the next-generation nuclear fusion reactors. This component will experience the highest thermal loads during the operation of a reactor since it directly faces the plasma. In recent years, after thorough analysis that followed a strategy of cost reduction, the ITER Organization decided to built a full-tunsgten divertor before the first nuclear campaigns. Therefore, tungsten will be used not only as a plasma-facing material (PFM) but also in structural applications. Tungsten, due to its the excellent thermo-physical properties fulfils the requirements of a PFM, however, its use in structural applications is compromised due to its inherent brittleness. One of the objectives of this phD thesis is therefore, to find a material with improved brittleness behaviour. The microstructural and mechanical characterisation of different tunsgten-based materials was performed. However, this is a challenging task because of the reduced laboratory-scale size of the specimens provided, their _ne microstructure and their brittleness. Consequently, many techniques are required to ensure an accurate measurement of all the mechanical and physical properties. Some of the applied methods have been widely used such as nanoindentation or three-point bending (TPB) tests. However, other methods were specifically developed and implemented during this work such as the measurement of the real fracture toughness of bulk-tunsgten alloys or the in situ fracture toughness measurements of very thin tungsten foils. Bulk-tunsgten materials with different compositions (W-1% Y2O3, W-2% V- 0.5% Y2O3, W-4% V-0.5% Y2O3, W-2% Ti-1% La2O3 and W-4% Ti-1% La2O3) were studied and compared with pure tungsten processed under the same conditions. These alloys, produced by a powder metallurgical route of mechanical alloying (MA) and hot isostatic pressing (HIP), were microstructural and mechanically characterised from 77 to 1473 K in air and under high vacuum conditions. Hardness, elastic modulus, flexural strength and fracture toughness for all of the alloys were measured in addition to other physical and mechanical properties. Finally, the fracture surfaces after the TPB tests were analysed to correlate the micromechanisms of failure with the macroscopic behaviour. The results reveal brittle mechanical behaviour in almost the entire temperature range for the alloys and micromechanisms of failure with no improvement in the ductile-brittle transition temperature (DBTT). To continue the search of a tungsten material with lowered DBTT, a preliminary study of pure tunsgten and 0.005 wt.% potassium (K)-doped tungsten foils was also performed. These foils were industrially produced by sintering and hot and cold rolling. After that, they were annealed from 1073 to 2673 K to analyse the evolution of the microstructural and mechanical properties with increasing annealing temperature. The results revealed the stabilisation of the tungsten grains with increasing annealing temperature in the potassium-doped tungsten foil. However, additional studies need to be performed to gain a better understanding of the microstructure and mechanical properties of these materials such as fracture toughness.

An engineering modification of the blade element momentum combination applicable in vertical descent flight :fundamentals and validation

An engineering modification of blade element/momentum theory is applied to describe the vertical autorotation of helicopter rotors. A full non‐linear aerodynamic model is considered for the airfoils, taking into account the dependence of lift and drag coefficients on both the angle of attack and the Reynolds number. The proposed model, which has been validated in previous work, has allowed the identification of different autorotation modes, which depend on the descent velocity and the twist of the rotor blades. These modes present different radial distributions of driven and driving blade regions, as well as different radial upwash/downwash patterns. The number of blade sections with zero tangential force, the existence of a downwash region in the rotor disk, the stability of the autorotation state, and the overall rotor autorotation efficiency, are all analyzed in terms of the flight velocity and the characteristics of the rotor. It is shown that, in vertical autorotation, larger blade twist leads to smaller values of descent velocity for a given thrust generated by the rotor in the autorotational state.

An engineering modification of the blade element momentum equation for vertical descent : an autoration case study

An engineering modification of blade element/momentum theory is applied to describe the vertical autorotation of helicopter rotors. A full non-linear aerodynamic model is considered for the airfoils, taking into account the dependence of lift and drag coefficients on both the angle of attack and the Reynolds number. The proposed model, which has been validated in previous work, has allowed the identification of different autorotation modes, which depend on the descent velocity and the twist of the rotor blades. These modes present different radial distributions of driven and driving blade regions, as well as different radial upwash/downwash patterns. The number of blade sections with zero tangential force, the existence of a downwash region in the rotor disk, the stability of the autorotation state, and the overall rotor autorotation efficiency, are all analyzed in terms of the flight velocity and the characteristics of the rotor. It is shown that, in vertical autorotation, larger blade twist leads to smaller values of descent velocity for a given thrust generated by the rotor in the autorotational state.

Covalent modification of the homeodomain-interacting protein kinase 2 (HIPK2) by the ubiquitin-like protein SUMO-1

Posttranslational modifications such as ubiquitination and phosphorylation play an important role in the regulation of cellular protein function. Homeodomain-interacting protein kinase 2 (HIPK2) is a member of the recently identified family of nuclear protein kinases that act as corepressors for homeodomain transcription factors. Here, we show that HIPK2 is regulated by a ubiquitin-like protein, SUMO-1. We demonstrate that HIPK2 localizes to nuclear speckles (dots) by means of a speckle-retention signal. This speckle-retention signal contains a domain that interacts with a mouse ubiquitin-like protein conjugating (E2) enzyme, mUBC9. In cultured cells, HIPK2 is covalently modified by SUMO-1, and the SUMO-1 modification of HIPK2 correlates with its localization to nuclear speckles (dots). Thus, our results provide firm evidence that the nuclear protein kinase HIPK2 can be covalently modified by SUMO-1, which directs its localization to nuclear speckles (dots).

Enhanced immunogenicity of HIV-1 vaccine construct by modification of the native peptide sequence

Viral proteins are not naturally selected for high affinity major histocompatibility complex (MHC) binding sequences; indeed, if there is any selection, it is likely to be negative in nature. Thus, one should be able to increase viral peptide binding to MHC in the rational design of synthetic peptide vaccines. The T1 helper peptide from the HIV-1 envelope protein was made more immunogenic for inducing T cell proliferation to the native sequence by replacing a residue that exerts an adverse influence on peptide binding to an MHC class II molecule. Mice immunized with vaccine constructs combining the more potent Th helper (Th) epitope with a cytotoxic T lymphocyte (CTL) determinant developed greatly enhanced CTL responses. Use of class II MHC-congenic mice confirmed that the enhancement of CTL response was due to class II-restricted help. Thus, enhanced T cell help is key for optimal induction of CTL, and, by modification of the native immunogen to increase binding to MHC, it is possible to develop second generation vaccine constructs that enhance both Th cell activation and CTL induction.

Conversion of cysteine to formylglycine: A protein modification in the endoplasmic reticulum

In sulfatases a Cα-formylglycine residue is found at a position where their cDNA sequences predict a cysteine residue. In multiple sulfatase deficiency, an inherited lysosomal storage disorder, catalytically inactive sulfatases are synthesized which retain the cysteine residue, indicating that the Cα-formylglycine residue is required for sulfatase activity. Using in vitro translation in the absence or presence of transport competent microsomes we found that newly synthesized sulfatase polypeptides carry a cysteine residue and that the oxidation of its thiol group to an aldehyde is catalyzed in the endoplasmic reticulum. A linear sequence of 16 residues surrounding the Cys-69 in arylsulfatase A is sufficient to direct the oxidation. This novel protein modification occurs after or at a late stage of cotranslational protein translocation into the endoplasmic reticulum when the polypeptide is not yet folded to its native structure.

Posttranscriptional modification of retroviral primers is required for late stages of DNA replication

During reverse transcription of retroviral RNA, synthesis of (−) strand DNA is primed by a cellular tRNA that anneals to an 18-nt primer binding site within the 5′ long terminal repeat. For (+) strand synthesis using a (−) strand DNA template linked to the tRNA primer, only the first 18 nt of tRNA are replicated to regenerate the primer binding site, creating the (+) strand strong stop DNA intermediate and providing a 3′ terminus capable of strand transfer and further elongation. On model HIV templates that approximate the (−) strand linked to natural modified or synthetic unmodified tRNA3Lys, we find that a (+) strand strong stop intermediate of the proper length is generated only on templates containing the natural, modified tRNA3Lys, suggesting that a posttranscriptional modification provides the termination signal. In the presence of a recipient template, synthesis after strand transfer occurs only from intermediates generated from templates containing modified tRNA3Lys. Reverse transcriptase from Moloney murine leukemia virus and avian myoblastosis virus shows the same requirement for a modified tRNA3Lys template. Because all retroviral tRNA primers contain the same 1-methyl-A58 modification, our results suggest that 1-methyl-A58 is generally required for termination of replication 18 nt into the tRNA sequence, generating the (+) strand intermediate, strand transfer, and subsequent synthesis of the entire (+) strand. The possibility that the host methyl transferase responsible for methylating A58 may provide a target for HIV chemotherapy is discussed.