Rustless translation

ATP binding cassette proteins are a large and diverse family of molecular machines and include transmembrane transporter, chromosome maintenance and DNA repair proteins, and translation factors. However, the function of the ABCE1, the only member of subfamily E of ABC proteins, remained mysterious for over a decade, even though it is perhaps the most conserved ABC protein in eukaryotes and archaea. Recent results have now identified ABCE1 as the ribosome-recycling factor of eukaryotes and archaea. Thus, two iron-sulfur clusters - the hallmark feature of ABCE1 - help catalyze an integral step of the translational cycle at the core of the protein synthesis machinery

Characterization of the longitudinal HIV-1 quasispecies evolution in HIV-1 infected individuals co-infected with Mycobacterium tuberculosis

One of the earliest and most striking observations made about HIV is the extensive genetic variation that the virus has within individual hosts, particularly in the hypervariable regions of the env gene which is divided into 5 variable regions (V1-V5) and 5 more constant (C1-C5) regions. HIV evolves at any time over the course of an individual’s infection and infected individuals harbours a population of genetically related but non-identical viruses that are under constant change and ready to adapt to changes in their environment. These genetically heterogeneous populations of closely related genomes are called quasispecies [65]. Tuberculosis or tubercle forming disease is an acute and/or chronic bacterial infection that primarily attacks the lungs, but which may also affect the kidneys, bones, lymph nodes, and brain. The disease is caused by Mycobacterium tuberculosis (MTB), a slow growing rod-shaped, acid fast bacterium. It is transmitted from person to person through inhalation of bacteria-carrying air droplets. Worldwide, one person out of three is infected with Mycobacterium tuberculosis – two billion people in total. TB currently holds the seventh place in the global ranking of causes of death [73]. In 2008, there were an estimated 9.4 (range, 8.9–9.9 million) million incident cases (equivalent to 139 cases per 100 000 population) of TB globally [75]. A complex biological interplay occurs between M. tuberculosis and HIV in coinfected host that results in the worsening of both pathologies. HIV promotes progression of M. tuberculosis either by endogenous reactivation or exogenous reinfection [77, 78] and, the course of HIV-1 infection is accelerated subsequent to the development of TB [80]. Active TB is associated with an increase in intra-patient HIV-1 diversity both systemically and at the infected lung sites [64,122]. The sustainability or reversal of the HIV-1 quasispecies heterogeneity after TB treatment is not known. Tetanus toxoid vaccinated HIV-1 infected patients developed a transient increase in HIV-1 heterogeneity which was reversed after few weeks [121]. Emergence of a heterogeneous HIV-1 population within a patient may be one of the mechanisms to escape strong immune or drug pressure [65,128]. The existence of better fitting and/or immune escape HIV-variants can lead to an increase in HIV-1 replication [129,130]. It might be that TB favourably selected HIV-1 variants which are sources for consistent HIV-1 replication. Understanding the mechanisms underlying the impacts of TB on HIV-1 is essential for the development of effective measures to reduce TB related morbidity and mortality in HIV-1 infected individuals. In the present study we studied whether the increase in HIV-1 quasispecies diversity during active TB is reversed or preserved throughout the course of antituberculous chemotherapy. For this purpose Two time point HIV-1 quasispecies were evaluated by comparing HIV-1 infected patients with active tuberculosis (HIV-1/TB) and HIV-1 infected patients without tuberculosis (HIV-1/non TB). Plasma samples were obtained from the Frankfurt HIV cohort and HIV-1 RNA was isolated. C2V5 env was amplified by PCR and molecular cloning was performed. Eight to twenty five clones were sequenced from each patient. Various phylogenetic analyses were performed including tree inferences, intra-patient viral diversity and divergence, selective pressure, co-receptor usage prediction and two time point identity of quasispecies comparison using Mantel’s test. We found out from this study that: 1) Active TB sustains HIV-1 quasispecies diversity for longer period 2. Active TB increases the rate of HIV-1 divergence 3) TB might slow down evolution of X4 variants And we concluded that active TB has an impact on HIV-1 viral diversity and divergence over time. The influence of active TB on longitudinal evolution of HIV- 1 may be predominant for R5 viruses. The use of CCR5-coreceptor inhibitors for HIV-1/TB patients as therapeutic approach needs further investigation.Eine der ersten und überraschenden Beobachtungen, welche bei der Analyse des HI-Virus gemacht wurden ist seine ausgeprägte Genetische Variabilität besonders die hypervariable Region des env Genes betreffen. Dieses wird in 5 variable Regionen (V1-V5) sowie 5 stärker konservierte Regionen (C1-C5) unterteilt. HIV wandelt sich zu jedem Zeitpunkt im Verlauf der Infektion und jedes infizierte Individuum ist Träger einer Population von genetisch verwandten jedoch nicht identischen Viren, welche sich kontinuierlich verändern und an die Erfordernisse innerhalb der Umgebung anpassen. Diese genetisch heterogenen, jedoch eng verwandten Populationen werden Quasispecies genannt. Tuberkulose ist eine mykobakterielle Infektion, welche sowohl akute als auch chronische Verläufe zeigt. Neben den Lungen als primärem Manifestationsort können auch die Nieren, Knochen und andere Organe befallen sein. Eine von drei Personen weltweit ist mit Mycobacterium tuberculosis infiziert, insgesamt 2 Milliarden Menschen. In HIV/TB Co-Inifzierten Menschen entsteht ein komplexes Zusammenspiel zwischen HIV und M. tuberculosis, welches zu einer Verschlechterung beider Krankheitsbilder führt. HIV führt durch endogene Rekativierung oder exogene Re-Infektion zu einer Progression der Tuberkulose, welche im weiteren Verlauf die Krankheitsprogression von HIV beschleunigt. Sowohl Morbidität als auch Mortalität sind in HIV-1/TB Co-Infizierten Menschen erhöht. Aktive Lungentuberkulose und Miliartuberkulose gehen mit dem Anstieg der Diversifität der HIV Viren innerhalb eines Wirtes einher. Wie lange diese erhöhte Heterogenität der HIV Quasispecies nach der erfolgreichen Behandlung einer Tuberkulose bestehen bleibt ist bisher noch unklar. Das Verständnis des dem Zusammenspiel von HIV und TB zugrundeliegenden Mechanismus ist essentiell für die Entwicklung von effektiven Massnahmen zur Senkung der Morbidität und Mortalität in HIV/TB Co-infizierten Menschen. Die gegenwärtige Forschungsarbeit folgte daher der Frage, ob wärend einer aktiven TB Infektion eine Zunahme der Diversität der HIV-1 Quasispecies zu beobachten ist und ob diese Diversität während einer TB Therapie erhalten bleibt oder sich zurück bildet. Hierfür wurden die HIV-1 Quasispecies zu zwei Zeitpunkten untersucht, wobei Proben von HIV-1 infizierten Patienten mit aktiver Tuberkulose (HIV-1/TB) und HIV infizierte Patienten ohne Tuberkulose (HIV-1/non TB) verglichen wurden. Aus Plasmaproben der Frankfurter HIV Cohorte wurde HIV-1 RNA isoliert. C2V5 env wurde durch PCR amplifiziert und molekular cloniert. Acht bis fünfundzwanzig Clone wurden für jeden Patienten sequenziert. Mehrere phylogenetische Analysen wurden durchgeführt, welche tree inferences, Intra-Patienten- und virale Diversität und Divergenz, Selektionsdruckanalysen, Vorhersage der Co-Rezeptornutzung sowie Zweipunktanalysen der Identität von Quasispecies mit Hilfe des Mantel’s Test miteinschlossen. Die Analysen ergaben die folgenden Ergebnisse: 1) Eine aktive TB erhält die Diversität von HIV-1 Quasispecies über einen längeren Zeitraum. 2. Eine aktive TB verstärkt die HIV -1 Divergenz 3) TB könnte zu einer langsameren Evolution von X4 Varianten führen. Schlussfolgerung: eine aktive TB beeinflusst die Entwicklung der viralen Diversität und Divergenz von HIV-1 im Verlauf der Krankheit. Der Einfluss der aktiven TB auf die longitudinale Evolution von HIV-1 könnte insbesondere R5 Viren betreffen. Der Einsatz von CCR5-Corezeptor Inhibitoren in HIV-1/TB coinifizerten Patienten sollte daher in Langzeitstudien untersucht werden

Aptamer-based therapeutics and their potential in radiopharmaceutical design

Aptamers, short, single stranded oligonucleotide entities, have been developed in the past 15 years against a plethora of targets and for a variety of applications. These range from inhibition of receptors and enzymes to the identification of small molecules in sensor applications, and from the development of targeted therapeutic to the design of novel diagnostic and imaging agents. Furthermore, aptamers have been designed for targets that cover a wide range of diseases, from HIV to tropical diseases, cancer and inflammation. Their easy development and flexibility of use and manipulation, offers further potential. In this paper we review their selection and consider some of the recent applications of aptamers in the design of radiopharmaceuticals for the targeted radiotherapy and medical imaging of disease

Infectious agents in atherosclerotic cardiovascular diseases through oxidative stress

Accumulating evidence demonstrates that vascular oxidative stress is a critical feature of atherosclerotic process, potentially triggered by several infectious agents that are considered as risk co-factors for the atherosclerotic cardiovascular diseases (CVDs). C. pneumoniae has been shown to upregulate multiple enzymatic systems capable of producing reactive oxygen species (ROS) such as NADPH oxidase (NOX) and cyclooxygenase in vascular endothelial cells, NOX and cytochrome c oxidase in macrophages as well as nitric oxide synthase and lipoxygenase in platelets contributing to both early and late stages of atherosclerosis. P. gingivalis seems to be markedly involved in the atherosclerotic process as compared to A. actinomycetemcomitans contributing to LDL oxidation and foam cell formation. Particularly interesting is the evidence describing the NLRP3 inflammasome activation as a new molecular mechanism underlying P. gingivalis-induced oxidative stress and inflammation. Amongst viral agents, immunodeficiency virus-1 and hepatitis C virus seem to have a major role in promoting ROS production, contributing, hence, to the early stages of atherosclerosis including endothelial dysfunction and LDL oxidation. In conclusion, oxidative mechanisms activated by several infectious agents during the atherosclerotic process underlying CVDs are very complex and not well-known, remaining, thus, an attractive target for future research

Human cytochrome P450 2B6 genetic variability in Botswana: a case of haplotype diversity and convergent phenotypes

Identification of inter-individual variability for drug metabolism through cytochrome P450 2B6 (CYP2B6) enzyme is important for understanding the di erences in clinical responses to malaria and HIV. This study evaluates the distribution of CYP2B6 alleles, haplotypes and inferred metabolic phenotypes among subjects with di erent ethnicity in Botswana. A total of 570 subjects were analyzed for CYP2B6 polymorphisms at position 516G>T (rs3745274), 785A>G (rs2279343) and 983T>C (rs28399499). Samples were collected in three districts of Botswana where the population belongs to Bantu (Serowe/Palapye and Chobe) and San-related (Ghanzi) ethnicity. The three districts showed di erent haplotype composition according to the ethnic background but similar metabolic inferred phenotypes, with 59.12%, 34.56%, 2.10% and 4.21% of the subjects having, respectively, an extensive, intermediate, slow and rapid metabolic pro le. The results hint at the possibility of a convergent adaptation of detoxifying metabolic phenotypes despite a di erent haplotype structure due to the di erent genetic background. The main implication is that, while there is substantial homogeneity of metabolic inferred phenotypes among the country, the response to drugs metabolized via CYP2B6 could be individually associated to an increased risk of treatment failure and toxicity. These are important facts since Botswana is facing malaria elimination and a very high HIV prevalence

A Unifying Scenario on the Origin and Evolution of Cellular and Viral Domains

The cellular theory on the nature of life has been one of the first major advancements in biology. Viruses, however, are the most abundant life forms, and their exclusion from mainstream biology and the Tree of Life (TOL) is a major paradox in biology. This article presents a broad, unifying scenario on the origin and evolution of cellular and viral domains that challenges the conventional views about the history of life and supports a TOL that includes viruses. Co-evolution of viruses and their host cells has led to some of the most remarkable developments and transitions in the evolution of life, including the origin of non-coding DNA as a genomic protective device against viral insertion damage. However, one of the major fundamental evolutionary developments driven by viruses was probably the origin of cellular domains - Bacteria, Archaea and Eukarya - from the Last Universal Common Ancestor (LUCA) lineage, by evolving anti-fusion mechanisms. Consistent with a novel fusion/fission model for the population mode of evolution of LUCA, this paper presents a “cell-like world” model for the origin of life. According to this model the evolution of coupled replication, transcription and translation system (RT&T) occurred within non-living cell-like compartments (CCs). In this model, the ancestral ribosome originated as template-based RNA synthesizing machinery. The origin of the cellular genome as a centralized unit for storage and replication of genetic information within the CCs facilitated the evolution of the ancestral ribosome into a powerful translation machinery - the modern ribosome. After several hundred millions of years of providing an enclosed environment and fusion/fission based exchanges necessary for the population mode of evolution of the basic metabolism and the RT&T, the CCs evolved into the first living entities on earth - the LUCA lineage. The paper concludes with a proposal for a TOL that integrates the co-evolution of cellular and viral domains. This is one of a series of three articles that present a unifying scenario on the origin and evolution of viral and cellular domains, including the origin of life, which has significant t bio-medical implications and could lead to a significant paradigm shift in biology

Co-Catalytic Metallopeptidases as Pharmaceutical Targets

Understanding the reaction mechanism of co-catalytic metallopeptidases provides a starting point for the design and synthesis of new molecules that can be screened as potential pharmaceuticals. Many of the enzymes that contain co-catalytic metallo-active sites play important roles in cellular processes such as tissue repair, protein maturation, hormone level regulation, cell-cycle control and protein degradation. Therefore, these enzymes play central roles in several disease states including cancer, HIV, stroke, diabetes, bacterial infections, neurological processes, schizophrenia, seizure disorders, and amyotrophic lateral sclerosis. The mechanism of AAP, an aminopeptidase from Aeromonas proteolytica, is one of the best-characterized examples of a metallopeptidase containing a co-catalytic metallo-active site, although this enzyme is not a specific pharmaceutical target at this time. As a large majority of co-catalytic metallopeptidases contain active sites that are nearly identical to the one observed in AAP, the major steps of their catalytic mechanisms are likely to be very similar. With this in mind, it is possible to propose a general catalytic mechanism for the hydrolysis of amino acid substrates

Adaptive HIV-1 evolutionary trajectories are constrained by protein stability

Despite the use of combination antiretroviral drugs for the treatment of HIV-1 infection, the emergence of drug resistance remains a problem. Resistance may be conferred either by a single mutation or a concerted set of mutations. The involvement of multiple mutations can arise due to interactions between sites in the amino acid sequence as a consequence of the need to maintain protein structure. To better understand the nature of such epistatic interactions, we reconstructed the ancestral sequences of HIV-1's Pol protein, and traced the evolutionary trajectories leading to mutations associated with drug resistance. Using contemporary and ancestral sequences we modelled the effects of mutations (i.e. amino acid replacements) on protein structure to understand the functional effects of residue changes. Although the majority of resistance-associated sequences tend to destabilise the protein structure, we find there is a general tendency for protein stability to decrease across HIV-1's evolutionary history. That a similar pattern is observed in the non-drug resistance lineages indicates that non-resistant mutations, for example, associated with escape from the immune response, also impacts on protein stability. Maintenance of optimal protein structure therefore represents a major constraining factor to the evolution of HIV-1

Architecturally diverse proteins converge on an analogous mechanism to inactivate Uracil-DNA glycosylase

Uracil-DNA glycosylase (UDG) compromises the replication strategies of diverse viruses from unrelated lineages. Virally encoded proteins therefore exist to limit, inhibit or target UDG activity for proteolysis. Viral proteins targeting UDG, such as the bacteriophage proteins ugi, and p56, and the HIV-1 protein Vpr, share no sequence similarity, and are not structurally homologous. Such diversity has hindered identification of known or expected UDG-inhibitory activities in other genomes. The structural basis for UDG inhibition by ugi is well characterized; yet, paradoxically, the structure of the unbound p56 protein is enigmatically unrevealing of its mechanism. To resolve this conundrum, we determined the structure of a p56 dimer bound to UDG. A helix from one of the subunits of p56 occupies the UDG DNA-binding cleft, whereas the dimer interface forms a hydrophobic box to trap a mechanistically important UDG residue. Surprisingly, these p56 inhibitory elements are unexpectedly analogous to features used by ugi despite profound architectural disparity. Contacts from B-DNA to UDG are mimicked by residues of the p56 helix, echoing the role of ugi’s inhibitory beta strand. Using mutagenesis, we propose that DNA mimicry by p56 is a targeting and specificity mechanism supporting tight inhibition via hydrophobic sequestration