Treatment of HIV-related primary central nervous system lymphoma with azt high dose, HAART, interleukin-2 and foscarnet in three patients

<p>Abstract</p> <p>Purpose</p> <p>Combined immunomodulatory and antiviral treatment was administered to three patients with newly diagnosed HIV-associated primary central nervous system lymphoma (PCNSL) in an attempt to improve outcomes.</p> <p>Patients and methods</p> <p>Three patients from our institution who were recently diagnosed with HIV-associated PCNSL received intravenous azidothymidine (AZT) 1.6 gr. bid for two weeks, followed by oral AZT 250 mg bid from day 15. In addition, complementary highly active antiretroviral therapy (HAART) with a second nucleoside reverse transcriptase inhibitor (NRTI) plus one protease inhibitor (PI) and interleukin 2 (IL-2) subcutaneously 2 million units twice daily (bid) plus foscarnet 90 mg/kg bid were administered on days 1-14. One patient received anti-EpsteinBarr virus (EBV)-maintenance therapy with ganciclovir, followed by cidofovir <abbrgrp><abbr bid="B1">1</abbr></abbrgrp>.</p> <p>Results</p> <p>All patients experienced progressive disease while on induction therapy, and switched early to whole-brain radiation therapy (WBRT) as second linetreatment. No grade 3 or 4 toxicities were observed. Two patients died on days 50 and 166 respectively due to progressive disease. The third patient with histologically proven lymphoproliferation and only suspected PCNSL remained alive at 53 months. He was on HAART and remained clinically and neurologically stable.</p> <p>Conclusion</p> <p>Although IL-2, HAART, high-dose AZT and foscarnet are used for other HIV-related conditions, they did not demonstrate benefit in lymphoma remission for 2 HIVassociated PCNSL patients. The third patient went into delayed remission after additional radiotherapy and was in good clinical and neurological health status over 53 months after diagnosis.</p

A new MR-SAD algorithm for the automatic building of protein models from low-resolution X-ray data and a poor starting model

Determining macromolecular structures from X-ray data with resolution worse than 3 Å remains a challenge. Even if a related starting model is available, its incompleteness or its bias together with a low observation-to-parameter ratio can render the process unsuccessful or very time-consuming. Yet, many biologically important macromolecules, especially large macromolecular assemblies, membrane proteins and receptors, tend to provide crystals that diffract to low resolution. A new algorithm to tackle this problem is presented that uses a multivariate function to simultaneously exploit information from both an initial partial model and low-resolution single-wavelength anomalous diffraction data. The new approach has been used for six challenging structure determinations, including the crystal structures of membrane proteins and macromolecular complexes that have evaded experts using other methods, and large structures from a 3.0 Å resolution F1-ATPase data set and a 4.5 Å resolution SecYEG–SecA complex data set. All of the models were automatically built by the method to Rfree values of between 28.9 and 39.9% and were free from the initial model bias

A simple and versatile microfluidic device for efficient biomacromolecule crystallization and structural analysis by serial crystallography

Determining optimal conditions for the production of well diffracting crystals is a key step in every biocrystallography project. Here, a microfluidic device is described that enables the production of crystals by counter-diffusion and their direct on-chip analysis by serial crystallography at room temperature. Nine ‘nonmodel’ and diverse biomacromolecules, including seven soluble proteins, a membrane protein and an RNA duplex, were crystallized and treated on-chip with a variety of standard techniques including micro-seeding, crystal soaking with ligands and crystal detection by fluorescence. Furthermore, the crystal structures of four proteins and an RNA were determined based on serial data collected on four synchrotron beamlines, demonstrating the general applicability of this multipurpose chip conceptThe following funding is acknowledged: Agence Nationale de la Recherche (contract No. ANR-11-LABX- 0057_MITOCROSS to Claude Sauter, Bernard Lorber; contract No. ANR-10-LABX-0036_NETRN to Claude Sauter, Bernard Lorber; contract No. ANR-13-BS07-0007-01 to Eric Girard, Sylvain Engilberge); Ministère des Affaires Etrangères (contract No. PROCOPE Hubert Curien to Claude Sauter, Mario Mörl); Deutsche Forschungsgemeinschaft (contract No. Mo 634/10-1 to Mario Mörl, Heike Betat); Université de Strasbourg [grant No. Initiative d’excellence (IDEX) to Claude Sauter, Raphaël de Wijn]; Centre National de la Recherche Scientifique (grant No. MRCT- 2012_PTI_UPR9002 to Claude Sauter)

The Metagenome-Derived Enzymes LipS and LipT Increase the Diversity of Known Lipases

Triacylglycerol lipases (EC 3.1.1.3) catalyze both hydrolysis and synthesis reactions with a broad spectrum of substrates rendering them especially suitable for many biotechnological applications. Most lipases used today originate from mesophilic organisms and are susceptible to thermal denaturation whereas only few possess high thermotolerance. Here, we report on the identification and characterization of two novel thermostable bacterial lipases identified by functional metagenomic screenings. Metagenomic libraries were constructed from enrichment cultures maintained at 65 to 75°C and screened resulting in the identification of initially 10 clones with lipolytic activities. Subsequently, two ORFs were identified encoding lipases, LipS and LipT. Comparative sequence analyses suggested that both enzymes are members of novel lipase families. LipS is a 30.2 kDa protein and revealed a half-life of 48 h at 70°C. The lipT gene encoded for a multimeric enzyme with a half-life of 3 h at 70°C. LipS had an optimum temperature at 70°C and LipT at 75°C. Both enzymes catalyzed hydrolysis of long-chain (C12 and C14) fatty acid esters and additionally hydrolyzed a number of industry-relevant substrates. LipS was highly specific for (R)-ibuprofen-phenyl ester with an enantiomeric excess (ee) of 99%. Furthermore, LipS was able to synthesize 1-propyl laurate and 1-tetradecyl myristate at 70°C with rates similar to those of the lipase CalB from Candida antarctica. LipS represents the first example of a thermostable metagenome-derived lipase with significant synthesis activities. Its X-ray structure was solved with a resolution of 1.99 Å revealing an unusually compact lid structure

MxCuBE: a synchrotron beamline control environment customized for macromolecular crystallography experiments

MxCuBE is a beamline control environment optimized for the needs of macromolecular crystallography. This paper describes the design of the software and the features that MxCuBE currently provides

Experimental procedure for the characterization of radiation damage in macromolecular crystals

A novel automatic procedure to determine the sensitivity of macromolecular crystals to radiation damage is presented. The information extracted from this procedure can be directly used for optimal planning of data collection or/and beamline calibration

The ubiquitin-associated (UBA) 1 domain of Schizosaccharomyces pombe Rhp23 is essential for the recognition of ubiquitin-proteasome system substrates both in vitro and in vivo

The ubiquitin-proteasome system is essential for maintaining a functional cell. Not only does it remove incorrectly folded proteins, it also regulates protein levels to ensure their appropriate spatial and temporal distribution. Proteins marked for degradation by the addition of Lys(48)-linked ubiquitin (Ub) chains are recognized by shuttle factors and transported to the 26 S proteasome. One of these shuttle factors, Schizosaccharomyces pombe Rhp23, has an unusual domain architecture. It comprises an N-terminal ubiquitin-like domain that can recognize the proteasome followed by two ubiquitin-associated (UBA) domains, termed UBA1 and UBA2, which can bind Ub. This architecture is conserved up to humans, suggesting that both domains are important for Rhp23 function. Such an extent of conservation raises the question as to why, in contrast to all other shuttle proteins, does Rhp23 require two UBA domains? We performed in vitro Ub binding assays using domain swap chimeric proteins and mutated domains in isolation as well as in the context of the full-length protein to reveal that the Ub binding properties of the UBA domains are context-dependent. In vivo, the internal Rhp23 UBA1 domain provides sufficient Ub recognition for the protein to function without UBA2

A new MR-SAD algorithm for the automatic building of protein models from low-resolution X-ray data and a poor starting model

Determining macromolecular structures from X-ray data with resolution worse than 3 Å remains a challenge. Even if a related starting model is available, its incompleteness or its bias together with a low observation-to-parameter ratio can render the process unsuccessful or very time-consuming. Yet, many biologically important macromolecules, especially large macromolecular assemblies, membrane proteins and receptors, tend to provide crystals that diffract to low resolution. A new algorithm to tackle this problem is presented that uses a multivariate function to simultaneously exploit information from both an initial partial model and low-resolution single-wavelength anomalous diffraction data. The new approach has been used for six challenging structure determinations, including the crystal structures of membrane proteins and macromolecular complexes that have evaded experts using other methods, and large structures from a 3.0 Å resolution F_1-ATPase data set and a 4.5 Å resolution SecYEG–SecA complex data set. All of the models were automatically built by the method to R_(free) values of between 28.9 and 39.9% and were free from the initial model bias

The \u3cem\u3eChlamydomonas\u3c/em\u3e Genome Reveals the Evolution of Key Animal and Plant Functions

Chlamydomonas reinhardtii is a unicellular green alga whose lineage diverged from land plants over 1 billion years ago. It is a model system for studying chloroplast-based photosynthesis, as well as the structure, assembly, and function of eukaryotic flagella (cilia), which were inherited from the common ancestor of plants and animals, but lost in land plants. We sequenced the ∼120-megabase nuclear genome of Chlamydomonas and performed comparative phylogenomic analyses, identifying genes encoding uncharacterized proteins that are likely associated with the function and biogenesis of chloroplasts or eukaryotic flagella. Analyses of the Chlamydomonas genome advance our understanding of the ancestral eukaryotic cell, reveal previously unknown genes associated with photosynthetic and flagellar functions, and establish links between ciliopathy and the composition and function of flagella