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Introduction: Despite improved treatment and access to care, adolescent AIDS deaths are decreasing more slowly than in any other age group. There is lack of longitudinal data around adolescent adherence and the dynamics of viraemia over time. We aimed to describe patterns of detectable viral load (VL) in a cohort of adolescents attending an ARV clinic in Cape Town, South Africa. Methods: We conducted a retrospective cohort study of all patients on antiretroviral therapy aged 10 to 19 years. Participants were included if they underwent at least two VL measurements and remained in care at the Groote Schuur Hospita HIV Clinic for at least 24 months between 2002 and 2016. The primary outcome was two consecutive HIV VL >100 copies/mL, in line with the lower limit of detection of assays in use over the follow-up period. Results and discussion: Of the 482 screened participants, 327 met inclusion criteria. Most participants had perinatally acquired HIV (n = 314; 96%), and 170 (52%) were males. Overall, there were 203 episodes of confirmed detectable VL involving 159 (49% (95% CI 43% to 54%)) participants during the follow-up period. Six participants had genotyped resistance to protease inhibitors. Four of these never suppressed, while two suppressed on salvage regimens. Total follow-up time was 1723 person years (PY), of which 880 (51%) were contributed by the 159 participants who experienced detectable VL. Overal time with detectable VL was 370 PY. This comprised 22% of total follow-up time, and 42% of the follow-up time contributed by those who experienced detectable VL. The rate of detectable VL was 11.8 (95% CI 10.3 to 13.5) episodes per 100 PY. The risk increased by 24% for each year of increasing age (Relative Risk 1.24 (95% CI 1.17 to 1.31); p < 0.0001). There was no sex difference with respect to duration (p = 0.4), prevalence (p = 0.46) and rate (p = 0.608) of detectable VL. Conclusions: Clinicians need to be alert to the high prevalence of detectable VL during adolescence so as to pre-empt it and act swiftly once it is diagnosed. This study helps to highlight the risk of detectable VL that is associated with increase in age as well the high proportion of time that poorly adherent adolescents spend in this state. Keywords: adolescents; adherence; viral suppression; ARV; Sub-Saharan Africa, 1 | INTRODUCTION AIDS-related deaths in adolescents are decreasing much slower than in all other age groups despite improved treatment and access to care [1,2]. HIV/AIDS is one of the [...]

Apple snails (Pomacea Perry, 1810) are successful invaders that cause ecological perturbations, economic losses, and medical issues. A peculiar trait of this snail is a high biological potential, related to the absence of predators of their eggs. Eggs show protease inhibitor (PI) activity, originally ascribed to PcOvo perivitellin in the apple snail Pomacea canaliculata (Lamarck, 1822) but absent in PmPV1, the orthologoue of PcOvo, in eggs of the apple snail Pomacea maculata Perry, 1810. As egg fluid diminishes rat growth rate, an antidigestive effect, similar to plant defenses against herbivory, was hypothesized. However, PI activity has not been characterized in apple snail eggs. Here we identify and partially characterize P. canaliculata egg PI and improve our knowledge of the quaternary structure and evolution of PcOvo. Through N-terminal, transcriptomic or proteomic sequencing, and biochemical validation, we identified a Kunitz-type and a Kazal-type inhibitor that, though at low concentration in the egg, exhibit strong PI activity against trypsin, chymotrypsin, elastase, and subtilisin. Additionally, we report three new subunits for the non-digestible storage protein PcOvo. They are likely products of ancient gene duplication, as their sequences exhibit moderate similarity (30%). To our knowledge, this is the first report of Kazal-type inhibition among invertebrate eggs. Inhibiting varied proteases, PI seems an efficient adaptive trait that limits predator's capacity to digest egg nutrients. Key words: protease inhibitor, invasive species, snail, Pomacea, predation defenses, egg defenses. Les ampullaires (Pomacea Perry, 1810) sont des envahisseurs prolifiques qui causent des perturbations ecologiques, des pertes economiques et des problemes medicaux. Un caractere particulier de ces escargots est leur potentiel biologique eleve associe a l'absence de predateurs de leurs oeufs. Les oeufs presentent une activite antiprotease attribuee a l'origine a la perivitelline PcOvo chez l'escargot-pomme Pomacea canaliculata (Lamarck, 1822), mais absente dans la PmPV1, l'orthologue de la PcOvo, dans les oeufs de Pomacea maculata Perry, 1810. Comme le liquide des oeufs reduit le taux de croissance de rats, la presence d'un effet antidigestif semblable a des mecanismes de defense des plantes contre l'herbivorie a ete postulee. L'activite antiprotease dans les oeufs d'ampullaires n'a toutefois pas ete caracterisee. Nous etablissons et caracterisons partiellement une activite antiprotease des oeufs de P. canaliculata et ameliorons les connaissances sur la structure quaternaire et l'evolution de la PcOvo. Nous utilisons le sequençage transcriptomique ou proteomique de l'extremite N-terminale et la validation biochimique pour etablir la presence d'un inhibiteur de type Kunitz et un autre de type Kazal qui, bien qu'en faibles concentrations dans les oeufs, presentent une forte activite antiprotease contre la trypsine, la chymotrypsine, l'elastase et la subtilisine. Nous faisons en outre etat de trois nouvelles sous-unites pour la proteine de reserve non digestible PcOvo, qui sont vraisemblablement des produits de duplications geniques anciennes, puisque leurs sequences presentent une similitude moderee (30 %). À notre connaissance, il s'agit du premier signalement d'une inhibition de type Kazal pour des oeufs d'invertebres. En inhibant des proteases diverses, l'activite antiprotease semble etre un caractere evolutif efficace pour limiter la capacite des predateurs a digerer les nutriments des oeufs. [Traduit par la Redaction] Mots-cles : antiprotease, espece envahissante, escargot, Pomacea, mecanismes antipredation, mecanismes de defense des oeufs., Introduction The apple snail Pomacea canaliculata (Lamarck, 1822) is an invasive species. Among the multiple characteristics that make this snail an effective invader stands its high biological potential, as the [...]

To link to full-text access for this article, visit this link: http://dx.doi.org/10.1016/j.ydbio.2008.08.030 Byline: Christoph Scherfer (a), Huaping Tang (b), Zakaria Kambris (a), Nouara Lhocine (a), Carl Hashimoto (d), Bruno Lemaitre (a)(c) Keywords: Melanization; Sclerotization; Pigmentation; Innate immunity; Crystal cells; Serine protease; Eclosion Abstract: In insects the enzyme phenoloxidase (PO) catalyzes melanin deposition at the wound site and around parasitoid eggs. Its proenzyme prophenoloxidase (proPO) is proteolytically cleaved to active phenoloxidase by a cascade consisting of serine proteases and inhibited by serpins. The Drosophila genome encodes 29 serpins, of which only two, Serpin-27A (Spn27A) and Necrotic, have been analyzed in detail. Using a genetic approach, we demonstrate that the so far uncharacterized Serpin-28D (Spn28D, CG7219) regulates the proPO cascade in both hemolymph and tracheal compartments. spn28D is the serpin gene most strongly induced upon injury. Inactivation of spn28D causes pupal lethality and a deregulated developmental PO activation leading to extensive melanization of tissues in contact with air and pigmentation defects of the adult cuticle. Our data also show that Spn28D regulates hemolymph PO activity in both larvae and adults at a different level than Spn27A. Our data support a model in which Spn28D confines PO availability by controlling its initial release, while Spn27A is rather limiting the melanization reaction to the wound site. This study further highlights the complexity of the proPO cascade that can be differentially regulated in different tissues during development. Author Affiliation: (a) Centre de Genetique Moleculaire (CGM), CNRS, 91198 Gif-sur-Yvette, France (b) Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, CT 06510, USA (c) Global Health Institute, EPFL, Station 15, CH-1015 Lausanne, Switzerland (d) Department of Cell Biology, Yale University School of Medicine, New Haven, CT 06520, USA Article History: Received 4 March 2008; Revised 9 July 2008; Accepted 21 August 2008

Introduction: Pre-treatment HIV drug resistance (PDR) is an increasing problem in sub-Saharan Africa. Children are an especially vulnerable population to develop PDR given that paediatric second-line treatment options are limited. Although monitoring of PDR is important, data on the paediatric prevalence in sub-Saharan Africa and its consequences for treatment outcomes are scarce. We designed a prospective paediatric cohort study to document the prevalence of PDR and its effect on subsequent treatment failure in Nigeria, the country with the second highest number of HIV-infected children in the world. Methods: HIV-1-infected children ≤ 12 years, who had not been exposed to drugs for the prevention of mother-to-child transmission (PMTCT), were enrolled between 2012 and 2013, and followed up for 24 months in Lagos, Nigeria. Pre- antiretroviral treatment (ART) population-based pol genotypic testing and six-monthly viral load (VL) testing were performed. Logistic regression analysis was used to assess the effect of PDR (World Health Organization (WHO) list for transmitted drug resistance) on subsequent treatment failure (two consecutive VL measurements >1000 cps/ml or death). Results: Of the total 82 PMTCT-naive children, 13 (15.9%) had PDR. All 13 children harboured non-nucleoside reverse transcriptase inhibitor (NNRTI) mutations, of whom seven also had nucleoside reverse transcriptase inhibitor resistance. After 24 months, 33% had experienced treatment failure. Treatment failure was associated with PDR and a higher log VL before treatment initiation (adjusted odds ratio (aOR) 7.53 (95%CI 1.61-35.15) and 2.85 (95%CI 1.04-7.78), respectively). Discussion: PDR was present in one out of six Nigerian children. These high numbers corroborate with recent findings in other African countries. The presence of PDR was relevant as it was the strongest predictor of first-line treatment failure. Conclusions: Our findings stress the importance of implementing fully active regimens in children living with HIV. This includes the implementation of protease inhibitor (PI)-based first-line ART, as is recommended by the WHO for all HIV-infected children < 3 years of age. Overcoming practical barriers to implement PI-based regimens is essential to ensure optimal treatment for HIV-infected children in sub-Saharan Africa. In countries where individual VL or resistance testing is not possible, more attention should be given to paediatric PDR surveys. Keywords: HIV-1; paediatric; HIV drug resistance; PMTCT; sub-Saharan Africa; genotypic resistance testing., Introduction Pre-treatment drug resistance (PDR) forms an increasing threat to the success of antiretroviral treatment (ART) programmes in sub-Saharan Africa, where individual resistance testing is not routinely available [1]. In [...]

To link to full-text access for this article, visit this link: http://dx.doi.org/10.1016/j.ydbio.2005.08.001 Byline: Ching-Hang Wong, C. Yan Cheng Keywords: Testis; Adherens junction; Blood-testis barrier; Ectoplasmic specialization; Mitogen-activated protein kinases; Spermatogenesis Abstract: Mitogen-activated protein kinases (MAPKs) are important regulators of many cellular processes. In mammalian testes, these kinases are involved in controlling cell division, differentiation, survival and death, and are therefore critical to spermatogenesis. Recent studies have also illustrated their involvement in junction restructuring in the seminiferous epithelium, especially at the ectoplasmic specialization (ES), a testis-specific adherens junction (AJ) type. ES contributes to the adhesion between Sertoli cells at the blood-testis barrier, as well as between Sertoli and developing spermatids (step 9 and beyond) at the adluminal compartment. MAPKs regulate AJ dynamics in the testis via their effects on the turnover of junction-associated protein complexes, the production of proteases and protease inhibitors, and the cytoskeleton structure. In this review, roles of the three major MAPK members, namely extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 MAPK, in ES dynamics are critically discussed. An integrated model of how these three MAPKs regulate adhesion function in the seminiferous epithelium is also presented. This model will serve as the framework for future investigation in the field. Author Affiliation: Population Council, 1230 York Avenue, New York, NY 10021, USA Article History: Received 17 May 2005; Revised 5 July 2005; Accepted 2 August 2005

Acetoacetanilide and 4'-chloroacetoacetanilide were trilithiated with excess lithium diisopropylamide and condensed with several aromatic esters, followed by neutralization, separate acid cyclization, and rearrangements. After C-acylation of trilithiated acetoacetanilides and cyclization to 4H-pyran-4-ones, these compounds underwent multistep rearrangements to 4-anilino-6-aryl-2H-pyran-2-ones. Key words: pyranones, trianions, Claisen-type condensations, rearrangements.

Results show that incorporation of beta-amino acids into the design of inhibitors of metalloendopeptidase EP 24.15 and EP 24.16 decreases the inhibitory rate to a low micromolar range. Data show that the inhibitors are resistant to cleavage by peptidase, suggesting the role of beta-amino acids in stabilizing enzyme inhibitors.

The hydrophilic peptidic inhibitor, ATBI, from Bacillus sp., competitively inhibits aspartic proteases and pepsin leading to conformational changes in the proteases. Amino acid substitutions, involving Asp, Glu, and Lys, totally abolishes the antiproteolytic activity, suggesting participation of two carboxyl and one amine group of ATBI in the inactivation process.

Research elucidates the structure of the wild-type human cysteine protease inhibitor cystatin A by comparing N-terminally truncated mutant version and wild-type with a variant containing methionine-65 to leucine substitution in cystatin A. Data indicate that the structure of the latter is a reliable structural model for cystatin A.

The beta-secondary and solvent deuterium kinetic isotope effects on catalysis by the Streptomyces R61 DD-peptidase were examined via comparisons with a structurally similar class C beta-lactamase. The results suggested that proton transfer is not involved in the acylation transition state of a thioester, at least for the P99 beta-lactamase. However, significant proton transfer may be involved in the deacylation transition states of both enzymes, which could be due to the rise of general acid/base catalysis.

Introduction: Response to antiretroviral therapy (ART) among individuals infected with HIV-2 is poorly described. We compared the immunological response among patients treated with three nucleoside reverse-transcriptase inhibitors (NRTIs) to boosted protease inhibitor (PI) and unboosted PI-based regimens in West Africa. Methods: This prospective cohort study enrolled treatment-naive HIV-2-infected patients within the International Epidemiological Databases to Evaluate AIDS collaboration in West Africa. We used mixed models to compare the CD4 count response to treatment over 12 months between regimens. Results: Of 422 HIV-2-infected patients, 285 (67.5%) were treated with a boosted PI-based regimen, 104 (24.6%) with an unboosted PI-based regimen and 33 (7.8%) with three NRTIs. Treatment groups were comparable with regard to gender (54.5% female) and median age at ART initiation (45.3 years; interquartile range 38.3 to 51.8).Treatment groups differed by clinical stage (21.2%, 16.8% and 17.3% at CDC Stage C or World Health Organization Stage IV for the triple NRTI, boosted PI and unboosted PI groups, respectively, p =0.02), median length of follow-up (12.9,17.7 and 44.0 months for the triple NRTI, the boosted PI and the unboosted PI groups, respectively, p Conclusions: In this observational study using African data, boosted PI-containing regimens had better immunological response compared to triple NRTI combinations and unboosted PI-based regimens at 12 months. A randomized clinical trial is still required to determine the best initial regimen for treating HIV-2 infected patients. Keywords: HIV-2; immunological response; antiretroviral treatment; linear mixed models; West Africa., Introduction Between one and two million people are estimated to be living with HIV-2 infection in West Africa [1], the region that is the epicentre of the HIV-2 epidemic, with [...]

HIV-protease inhibitors have led to substantial reductions in blood levels of HIV, but they do not eradicate the virus completely. Nevertheless, most symptomatic HIV patients with CD cell counts less than 500 should receive protease inhibitors in combination with nucleoside or non-nucleoside drugs such as zidovudine. HIV-protease inhibitors include the drugs indinavir, ritonavir, saquinavir and nelfinavir. These drugs inhibit a viral enzyme that allows the virus to reproduce. They interact with some other drugs used in HIV patients and the patient can also develop resistance to the drugs.

The serpins are a family of proteins that inhibit chymotrypsin-like serine proteinases, with an unusual mechanism involving a large conformational change known as the stressed [right arrow] relaxed (S [right arrow] R) transition. This article is a guide to the known serpin conformations and their biological significance.

The inhibiting effect of SH3 domain binding peptides on Lyn function in mast cell activation was determined through utilization of electroporating Lyn-specific SH3 domain binding peptides into adherent RBL cells. Results have shown that the binding of cellular interaction partners to Lyn SH3 domain inhibits the Lyn kinase activity and lessens IgE-receptor cross-linking-induced receptor phosphorylation and calcium signaling.

The kinetic role of an NS4A peptide in the catalytic activation of the NS3 protease domain was determined following the use of steady-state kinetic analysis. The specificity of the NS3-NS4A complex toward amino acid substitutions at the P1 and P1' positions of peptide substrates was also analyzed. Results have shown that NBS4A regulates NS3 protease activity by modifying the S' subsites.

A 155-kDa proteinase inhibitor, pacifastin, from plasma of the freshwater crayfish, Pacifastacus leniusculus, was found to be composed of two covalently linked subunits. The two subunits are encoded by two different mRNAs, which were cloned and sequenced. The heavy chain of pacifastin (105 kDa) is related to transferrins, containing three transferrin lobes, two of which seem to be active for iron binding. The light chain of pacifastin (44 kDa) is the inhibitory subunit, and has nine cysteine-rich inhibitory domains that are homologous to each other and to low molecular weight proteinase inhibitors isolated from the grasshopper, Locusta migratoria. The nine light chain domains and the Locusta inhibitors share a characteristic cysteine array (Cys-[Xaa.sub.9-12]-Cys-[Xaa.sub.2]-Cys-Xaa-Cys-[Xaa.sub.6-8] Cys-[Xaa.sub.4]-Cys) distinct from any described proteinase inhibitor family, suggesting that they constitute a new family of proteinase inhibitors. Pacifastin is the first known protein that has combined properties of a transferrin-like molecule and a proteinase inhibitor.

The proteasome is a multicatalytic protease complex that plays a key role in diverse cellular functions. The peptide vinyl sulfone, carboxybenzyl-leucyl-leucyl-leucine vinyl sulfone (Z-[L.sub.3]VS) covalently inhibits the trypsin-like, chymotrypsin-like and, unlike lactacystin, also the peptidyl-glutamyl peptidase activity in isolated proteasomes, and blocks their function in living cells. Although described as a class of mechanism-based inhibitors for cysteine proteases, the peptide vinyl sulfone Z-[L.sub.3]VS and a 125I-labeled nitrophenol derivative (125I-NIP-[L.sub.3]VS) covalently modify the active site threonine of the catalytic [Beta] subunits of the proteasome. Modification of Thermoplasma proteasomes demonstrates the requirement for a hydroxyl amino acid (threonine, serine) as nucleophile at the [Beta] subunit's N[H.sub.2] terminus. 125I-NIP-[L.sub.3]VS covalently modifies the HslV subunit of the Escherichia coli protease complex HslV/HslU, a reaction that requires ATP, and supports a catalytic mechanism shared with that of the eukaryotic proteasome.

A study was conducted to demonstrate the proteolytic regulation of polypeptides intracellularly secreted by the bacterial pathogen Listeria monocytogenes. Three forms of polypeptides, which mediate bacterial cell-to-cell spread, were identified. These include two phospholipases of the C class (PLC), namely, phosphatidylinositol-specific PLC (PI-PLC) and broad spectrum PLC (PC-PLC). The third polypeptide form is a metalloprotease, which proteolytically activates PC-PLC as an inactive precursor (proPC-PLC).

Saccharomyces cerevisiae vacuole inheritance requires two low molecular weight activities, LMA1 and LMA2. LMA1 is a heterodimer of thioredoxin and protease B inhibitor 2 [Mathematical Expression Omitted]. Here we show that the second low molecular weight activity (LMA2) is monomeric [Mathematical Expression Omitted]. Though [Mathematical Expression Omitted] was initially identified as a protease B inhibitor, this protease inhibitor activity is not related to its ability to promote vacuole fusion: (i) Low [M.sub.r] protease B inhibitors cannot substitute for LMA1 or LMA 2, (ii) LMA1 and LMA2 promote the fusion of vacuoles from a strain that has no protease B, (iii) low concentrations of LMA2 that fully inhibit protease B do not promote vacuole fusion, and (iv) LMA1, in which [Mathematical Expression Omitted] is complexed with thioredoxin, is far more active than [Mathematical Expression Omitted] in promoting vacuole fusion and far less active in inhibiting protease B. These studies establish a new function for [Mathematical Expression Omitted].

The inhibitory activities of the arrowhead proteinase inhibitors A and B are different despite sharing 91% homology. The double-headed inhibitors comprise 179 amino acid residues with three disulfide bridges. Proteinase post-processing may be used to cleave off the seven additional residues in the C-terminal which were discovered through sequence analysis of the cDNA sequences. The structure-function relationship of the inhibitors is analyzed.

Secretory leukocyte protease inhibitor (SLPI), a known epithelial cell-derived inhibitor of leukocyte serine proteases, is transcribed by lipopolysaccharide (LPS)-hyporesponsive cells, primary macrophages and polymorphonuclear leukocytes. It can be considered a phagocyte-derived LPS-induced LPS inhibitor due to its sources, functions and modes of regulation. It is one of the few genes whose expression in macrophages is induced by LPS and suppressed by interferon-gamma (IFN-gamma). IFN-gamma suppressed expression of SLPI and restored LPS responsiveness to SLPI-producing cells.

Cysteine proteases which include the papain, calpain, streptopain, picornain and clostripain families, are found in viruses, bacteria, protozoa, plants and mammals. The mechanism of action of these enzymes is yet unknown although it has been recognized that the ion pair mechanism accounts for their unusually high reactivity. Endogenous cysteine protease inhibitors are classified into stefin, cystatin and kininogen families. A common feature of the synthetic inhibitors is a peptide segment that is bound to a nucleophilic attackable/substitutable group which can react with the cysteine residue of the active site.

A comparison of the X-ray crystal structures of 12 substrate-based peptidic inhibitors bound in the active site of the aspartyl protease, HIV-1 protease, revealed that the inhibitor-binding modes of these inhibitors are notably similar, despite their structural differences and conformational flexibility. A bicyclic peptidomimetic inhibitor with macrocyclic components in constrained conformations preorganized for receptor-binding is then designed based upon the enzyme-bound structures of the substrate-derived, peptidic inhibitors. It is a consensus conformational mimic of the protease-bound inhibitor structures with superior properties to peptides.

Both the stepwise solid phase peptide synthesis technique and the native chemical ligation method were used in the total chemical synthesis of turkey ovomucolid third domain, (6-56)OMTKY3. Characterization of the resultant products showed that the two synthetic proteins are identical in function and structure to their recombinant counterpart. The two techniques also provided comparable yields and similar product quality.

The isolation, structural analysis, chemistry and biological activity of actinoplanic acid A and its more active congener actinoplanic acid B are presented. The results showed that both compounds are highly functionalized 30-carbon chain length polyketides with a terminating carboxylic acid group. Both acids inhibit farnesyl-protein transferases which are competitive with respect to farnesyl pyrophosphate and uncompetitive with the peptide substrate, Ras-CVIM.

The structural stability of chymotrypsin inhibitor 2's partly exposed minicore is indicated by 17 mutant proteins. Mutations of the Leu51, Val57, and Phe69 residues in the hydrophobic minicore destabilize the protein in a range of 0.3-4.8 kcal/mol. The structure of the minicore is stabilized by hydrophobic interaction alone. The hydrophobic changes at the protein surface fit the classical model of hydrophobic effect.