Organic nitrate chemistry and its implications for nitrogen budgets in an isoprene- and monoterpene-rich atmosphere: constraints from aircraft (SEAC4RS) and ground-based (SOAS) observations in the Southeast US.

Formation of organic nitrates (RONO2) during oxidation of biogenic volatile organic compounds (BVOCs: isoprene, monoterpenes) is a significant loss pathway for atmospheric nitrogen oxide radicals (NOx), but the chemistry of RONO2 formation and degradation remains uncertain. Here we implement a new BVOC oxidation mechanism (including updated isoprene chemistry, new monoterpene chemistry, and particle uptake of RONO2) in the GEOS-Chem global chemical transport model with ∼25 × 25 km2 resolution over North America. We evaluate the model using aircraft (SEAC4RS) and ground-based (SOAS) observations of NOx, BVOCs, and RONO2 from the Southeast US in summer 2013. The updated simulation successfully reproduces the concentrations of individual gas- and particle-phase RONO2 species measured during the campaigns. Gas-phase isoprene nitrates account for 25-50% of observed RONO2 in surface air, and we find that another 10% is contributed by gas-phase monoterpene nitrates. Observations in the free troposphere show an important contribution from long-lived nitrates derived from anthropogenic VOCs. During both campaigns, at least 10% of observed boundary layer RONO2 were in the particle phase. We find that aerosol uptake followed by hydrolysis to HNO3 accounts for 60% of simulated gas-phase RONO2 loss in the boundary layer. Other losses are 20% by photolysis to recycle NOx and 15% by dry deposition. RONO2 production accounts for 20% of the net regional NOx sink in the Southeast US in summer, limited by the spatial segregation between BVOC and NOx emissions. This segregation implies that RONO2 production will remain a minor sink for NOx in the Southeast US in the future even as NOx emissions continue to decline.

Aqueous-phase mechanism for secondary organic aerosol formation from isoprene: application to the Southeast United States and co-benefit of SO2 emission controls.

Isoprene emitted by vegetation is an important precursor of secondary organic aerosol (SOA), but the mechanism and yields are uncertain. Aerosol is prevailingly aqueous under the humid conditions typical of isoprene-emitting regions. Here we develop an aqueous-phase mechanism for isoprene SOA formation coupled to a detailed gas-phase isoprene oxidation scheme. The mechanism is based on aerosol reactive uptake coefficients (γ) for water-soluble isoprene oxidation products, including sensitivity to aerosol acidity and nucleophile concentrations. We apply this mechanism to simulation of aircraft (SEAC4RS) and ground-based (SOAS) observations over the Southeast US in summer 2013 using the GEOS-Chem chemical transport model. Emissions of nitrogen oxides (NOx ≡ NO + NO2) over the Southeast US are such that the peroxy radicals produced from isoprene oxidation (ISOPO2) react significantly with both NO (high-NOx pathway) and HO2 (low-NOx pathway), leading to different suites of isoprene SOA precursors. We find a mean SOA mass yield of 3.3 % from isoprene oxidation, consistent with the observed relationship of total fine organic aerosol (OA) and formaldehyde (a product of isoprene oxidation). Isoprene SOA production is mainly contributed by two immediate gas-phase precursors, isoprene epoxydiols (IEPOX, 58% of isoprene SOA) from the low-NOx pathway and glyoxal (28%) from both low- and high-NOx pathways. This speciation is consistent with observations of IEPOX SOA from SOAS and SEAC4RS. Observations show a strong relationship between IEPOX SOA and sulfate aerosol that we explain as due to the effect of sulfate on aerosol acidity and volume. Isoprene SOA concentrations increase as NOx emissions decrease (favoring the low-NOx pathway for isoprene oxidation), but decrease more strongly as SO2 emissions decrease (due to the effect of sulfate on aerosol acidity and volume). The US EPA projects 2013-2025 decreases in anthropogenic emissions of 34% for NOx (leading to 7% increase in isoprene SOA) and 48% for SO2 (35% decrease in isoprene SOA). Reducing SO2 emissions decreases sulfate and isoprene SOA by a similar magnitude, representing a factor of 2 co-benefit for PM2.5 from SO2 emission controls.

Beneficial effects of judo training on bone mineral density of high-school boys in Korea.

Bone mineralization is strongly stimulated by weight-bearing exercise during growth and development. Judo, an Olympic combat sport, is a well-known form of strenuous and weight-bearing physical activity. Therefore, the primary goal of this study was to determine the effects of Judo practice on the bone health of male high school students in Korea. The secondary goal of this study was to measure and compare the bone mineral density (BMD) of the hands of Judo players and sedentary control subjects. Thirty Judo players (JDP) and 30 sedentary high school boys (CON) voluntarily participated in the present study, and all of the sedentary control subjects were individually matched to the Judo players by body weight. BMD was determined by using dual-energy X-ray absorptiometry (Hologic, Bedford, MA, USA). The lumbar spine, femur and forearm BMD in the JDP group were significantly greater by 22.7%, 24.5%, and 18.3%, respectively, than those in the CON group. In addition, a significant difference in the CON group was observed between the dominant hand (DH) radius (0.710 ± 0.074 g/cm(2)) and the non-dominant hand (NDH) radius (0.683 ± 0.072 g/cm(2)), but this was not observed in the JDP group (DH = 0.819 ± 0.055 g/cm(2); NDH = 810 ± 0.066 g/cm(2)) (P < 0.05). Therefore, the results of this study suggest that Judo practice during the growth period significantly improves bone health in high school male students. In addition, it seems that Judo practice could eliminate the effect of increased BMD in the dominant hand.

Arginase induction by sodium phenylbutyrate in mouse tissues and human cell lines.

Hyperargininemia is a urea cycle disorder caused by mutations in the gene for arginase I (AI) resulting in elevated blood arginine and ammonia levels. Sodium phenylacetate and a precursor, sodium phenylbutyrate (NaPB) have been used to lower ammonia, conjugating glutamine to produce phenylacetylglutamine which is excreted in urine. The elevated arginine levels induce the second arginase (AII) in patient kidney and kidney tissue culture. It has been shown that NaPB increases expression of some target genes and we tested its effect on arginase induction. Eight 9-week old male mice fed on chow containing 7.5 g NaPB/kg rodent chow and drank water with 10 g NaPB/L, and four control mice had a normal diet. After one week all mice were sacrificed. The arginase specific activities for control and NaPB mice, respectively, were 38.2 and 59.4 U/mg in liver, 0.33 and 0.42 U/mg in kidney, and 0.29 and 1.19 U/mg in brain. Immunoprecipitation of arginase in each tissue with AI and AII antibodies showed the activity induced by NaPB is mostly AI. AII may also be induced in kidney. AI accounts for the fourfold increased activity in brain. In some cell lines, NaPB increased arginase activity up to fivefold depending on dose (1-5 mM) and exposure time (2-5 days); control and NaPB activities, respectively, are: erythroleukemia, HEL, 0.06 and 0.31 U/mg, and K562, 0.46 and 1.74 U/mg; embryonic kidney, HEK293, 1.98 and 3.58 U/mg; breast adenocarcinoma, MDA-MB-468, 1.11 and 4.06 U/mg; and prostate adenocarcinoma, PC-3, 0.55 and 3.20 U/mg. In MDA-MB-468 and HEK most, but not all, of the induced activity is AI. These studies suggest that NaPB may induce AI when used to treat urea cycle disorders. It is relatively less useful in AI deficiency, although it could have some effect in those patients with missense mutations.

Diffusion-weighted MRI and ADC mapping in FK506 neurotoxicity.

FK506 is a newly developed potent immunosuppressant for preventing rejection after organ transplantation. However, FK506 can induce central nervous system toxicity. Until now the pathogenic mechanism of FK506 neurotoxicity was unclear. We report the findings of diffusion-weighted MRI and apparent diffusion coefficient (ADC) mapping of a FK506 neurotoxicity patient who showed increased signal intensities in both parieto-occipital lobes on T(2) weighted images, diffusion-weighted images and ADC maps. These findings suggest that a vasogenic oedema rather than a cytotoxic oedema may play a pivotal role in FK506 neurotoxicity pathogenesis.

Efficient assembly of an HIV-1/MLV Gag-chimeric virus in murine cells.

In human cells infected by HIV type 1 (HIV-1), the viral Gag protein directs the assembly of nascent viral particles at the plasma membrane. In murine cells, HIV-1 Gag fails to reach the plasma membrane and instead forms nonfunctional intracellular aggregates. The viral determinants of this species incompatibility are previously undefined. To address this problem, we replaced a region of HIV-1 Gag known to direct its localization, the matrix (MA) domain, with functionally homologous regions from Moloney murine leukemia virus (MLV), a murine retrovirus. An HIV-1 clone carrying such a chimeric Gag protein, designated murine HIV (MHIV), assembled more efficiently than nonchimeric HIV-1 and restored plasma membrane localization of Gag in murine cells. Increased efficiency of viral assembly in murine cells was observed from MHIV constructs carrying MLV MA in place of HIV-1 MA. Efficient processing of the HIV-1 capsid protein from the chimeric Gag polyprotein and subsequent infectivity of MHIV required the presence of MLV p12 in addition to MLV MA. These findings strongly suggest that the HIV-1 MA domain of HIV-1 Gag is responsible for the assembly defect in mouse cells. Although these MHIV do not recruit native HIV-1 Env efficiently, they are capable of single-round infection when produced by high-efficiency transfection of human 293 cells and provided with an HIV-1 Env lacking its cytoplasmic tail. With further adaptation, this chimeric MHIV approach may provide the basis for creating an infectious mouse model for HIV/AIDS.

Side-chain repacking calculations for predicting structures and stabilities of heterodimeric coiled coils.

An important goal in biology is to predict from sequence data the high-resolution structures of proteins and the interactions that occur between them. In this paper, we describe a computational approach that can make these types of predictions for a series of coiled-coil dimers. Our method comprises a dual strategy that augments extensive conformational sampling with molecular mechanics minimization. To test the performance of the method, we designed six heterodimeric coiled coils with a range of stabilities and solved x-ray crystal structures for three of them. The stabilities and structures predicted by the calculations agree very well with experimental data: the average error in unfolding free energies is <1 kcal/mol, and nonhydrogen atoms in the predicted structures superimpose onto the experimental structures with rms deviations <0.7 A. We have also tested the method on a series of homodimers derived from vitellogenin-binding protein. The predicted relative stabilities of the homodimers show excellent agreement with previously published experimental measurements. A critical step in our procedure is to use energy minimization to relax side-chain geometries initially selected from a rotamer library. Our results show that computational methods can predict interaction specificities that are in good agreement with experimental data.

Design of potent inhibitors of HIV-1 entry from the gp41 N-peptide region.

The HIV-1 gp41 envelope glycoprotein promotes fusion of the virus and cell membranes through the formation of a trimer-of-hairpins structure, in which the amino- and carboxyl-terminal regions of the gp41 ectodomain are brought together. Synthetic peptides derived from these two regions (called N and C peptides, respectively) inhibit HIV-1 entry. In contrast to C peptides, which inhibit in the nanomolar range, N peptides are weak inhibitors with IC(50) values in the micromolar range. To test the hypothesis that the weak inhibition of N peptides results from their tendency to aggregate, we have constructed chimeric variants of the N-peptide region of gp41 in which soluble trimeric coiled coils are fused to portions of the gp41 N peptide. These molecules, which present the N peptide in a trimeric coiled-coil conformation, are remarkably more potent inhibitors than the N peptides themselves and likely target the carboxyl-terminal region of the gp41 ectodomain. The best inhibitors described here inhibit HIV-1 entry at nanomolar concentrations.

The trimer-of-hairpins motif in membrane fusion: Visna virus.

Structural studies of viral membrane fusion proteins suggest that a "trimer-of-hairpins" motif plays a critical role in the membrane fusion process of many enveloped viruses. In this motif, a coiled coil (formed by homotrimeric association of the N-terminal regions of the protein) is surrounded by three C-terminal regions that pack against the coiled coil in an oblique antiparallel manner. The resulting trimer-of-hairpins structure serves to bring the viral and cellular membranes together for fusion. learncoil-vmf, a computational program developed to recognize coiled coil-like regions that form the trimer-of-hairpins motif, predicts these regions in the membrane fusion protein of the Visna virus. Peptides corresponding to the computationally identified sequences were synthesized, and the soluble core of the Visna membrane fusion protein was reconstituted in solution. Its crystal structure at 1.5-A resolution demonstrates that a trimer-of-hairpins structure is formed. Remarkably, despite less than 23% sequence identity, the ectodomains in Visna and HIV-1 envelope glycoproteins show detailed structural conservation, especially within the area of a hydrophobic pocket in the central coiled coil currently being targeted for the development of new anti-HIV drugs.

A designed protein with packing between left-handed and right-handed helices.

A common motif in protein structures is the assembly of alpha-helices. Natural alpha-helical assemblies, such as helical bundles and coiled coils, consist of multiple right-handed alpha-helices. Here we design a protein complex containing both left-handed and right-handed helices, with peptides of D- and L-amino acids, respectively. The two peptides, D-Acid and L-Base, feature hydrophobic heptad repeats and are designed to pack against each other in a "knobs-into-holes" manner. In solution, the peptides form a stable, helical heterotetramer with tight packing in the most solvent-protected core. This motif may be useful for designing protease-resistant, helical D-peptide ligands against biological protein targets.

A multiethnic study of Delta32ccr5 and ccr2b-V64I allele distribution in four Los Angeles populations.

Mutant alleles of the chemokine receptors CCR5 and CCR2 affect the susceptibility to HIV infection as well as the rate of disease progression. In this article the authors report the results of a survey for presence of the common Delta32ccr5 and ccr2b-V64I mutant alleles in 472 individuals of a multiethnic cohort. Hispanic Americans had the highest observed frequency of the Delta32ccr5 allele (3.57%), whereas African Americans had a lower frequency (1.55%). The mutant allele was absent in Asian Americans and Native Americans. Thus, the Delta32ccr5 allele segregates in populations with a significant white admixture and is rare in genetically distant non-European groups. Native Americans had the highest occurrence of the ccr2b-V64I allele (31.13%), whereas African Americans, Asian Americans, and Hispanic Americans had much lower frequencies (14.36%, 11.94%, and 14.37% respectively). This mutation is probably an ancient one, occurring before the migration of the ancestors of Native Americans across the Bering Straits to the Americas. The twofold greater frequency of ccr2b-V64I in modern Native Americans probably reflects a founder effect. The observed population differences in Delta32ccr5 and ccr2b-V64I frequencies, considered together with their documented effects on sensitivity to HIV infection and rate of disease progression, have implications for HIV transmission patterns in the United States, as well as for AIDS prediction, monitoring, and treatment.

Mechanisms of viral membrane fusion and its inhibition.

Viral envelope glycoproteins promote viral infection by mediating the fusion of the viral membrane with the host-cell membrane. Structural and biochemical studies of two viral glycoproteins, influenza hemagglutinin and HIV-1 envelope protein, have led to a common model for viral entry. The fusion mechanism involves a transient conformational species that can be targeted by therapeutic strategies. This mechanism of infectivity is likely utilized by a wide variety of enveloped viruses for which similar therapeutic interventions should be possible.

Buried polar residues in coiled-coil interfaces.

Coiled coils, estimated to constitute 3-5% of the encoded residues in most genomes, are characterized by a heptad repeat, (abcdefg)(n), where the buried a and d positions form the interface between multiple alpha-helices. Although generally hydrophobic, a substantial fraction ( approximately 20%) of these a- and d-position residues are polar or charged. We constructed variants of the well-characterized coiled coil GCN4-p1 with a single polar residue (Asn, Gln, Ser, or Thr) at either an a or a d position. The stability and oligomeric specificity of each variant were measured, and crystal structures of coiled-coil trimers with threonine or serine at either an a or a d position were determined. The structures show how single polar residues in the interface affect not only local packing, but also overall coiled-coil geometry as seen by changes in the Crick supercoil parameters and core cavity volumes.

Radiofrequency sacroiliac joint denervation for sacroiliac syndrome.

BACKGROUND AND OBJECTIVES: Radiofrequency (RF) denervation of the sacroiliac (SI) joint has been advocated for the treatment of sacroiliac syndrome, yet no clinical studies or case series support its use. METHODS: We report the results of a consecutive series of 50 SI joint RF denervations performed in 33 patients with sacroiliac syndrome. All patients underwent diagnostic SI joint injections with local anesthetic before denervation. Changes in visual analog pain scores (VAS), pain diagrams, physical examination (palpation tenderness over the joint, myofascial trigger points overlying the joint, SI joint pain provocation tests, and range of motion of the lumbar spine), and opioid use were assessed pre- and postdenervation. RESULTS: The criteria for successful RF denervation were at least a 50% decrease in VAS for a period of at least 6 months; 36.4% of patients (12 of 33) met these criteria. Failure of denervation correlated with the presence of disability determination and pain on lateral flexion to the affected side. The average duration of pain relief was 12.0 +/- 1.2 months in responders versus 0.9 +/- 0.2 months in nonresponders (P < or = 0.0001). A positive response was associated with an atraumatic inciting event. Successful denervation was associated with a change in the pain diagram and a reduction in the pattern of referred pain, a normalization of SI joint pain provocation tests, and a reduction in the use of opioids. CONCLUSIONS: This study suggests that RF denervation of the SI joint can significantly reduce pain in selected patients with sacroiliac syndrome for a protracted time period. Moreover, certain abnormal physical findings (i.e., SI joint pain provocation tests) revert to normal for the duration of the analgesia.

Protein design of an HIV-1 entry inhibitor.

Human immunodeficiency virus type-1 (HIV-1) membrane fusion is promoted by the formation of a trimer-of-hairpins structure that brings the amino- and carboxyl-terminal regions of the gp41 envelope glycoprotein ectodomain into close proximity. Peptides derived from the carboxyl-terminal region (called C-peptides) potently inhibit HIV-1 entry by binding to the gp41 amino-terminal region. To test the converse of this inhibitory strategy, we designed a small protein, denoted 5-Helix, that binds the C-peptide region of gp41. The 5-Helix protein displays potent (nanomolar) inhibitory activity against diverse HIV-1 variants and may serve as the basis for a new class of antiviral agents. The inhibitory activity of 5-Helix also suggests a strategy for generating an HIV-1 neutralizing antibody response that targets the carboxyl-terminal region of the gp41 ectodomain.

Savinin, a lignan from Pterocarpus santalinus inhibits tumor necrosis factor-alpha production and T cell proliferation.

Two lignans were isolated from the heartwood of Pterocarpus santalinus by activity-guided fractionation and investigated for their biological properties and molecular mechanism of action. On the basis of their spectroscopic data, these compounds were identified as savinin (1) and calocedrin (2), dibenzyl butyrolactone-type lignan compounds having an alpha-arylidene gamma-lactone structure. These lignans significantly inhibited tumor necrosis factor (TNF)-a production in lipopolysaccharide (LPS)-stimulated RAW264.7 cells, and T cell proliferation elicited by concanavalin (Con A), without displaying cytotoxicity. The molecular inhibitory mechanism of compound 1 was confirmed to be mediated by the non-polar butyrolactone ring, according to a structure-relationship study with structurally related and unrelated compounds, such as arctigenin (a dibenzyl butyrolactone type lignan), eudesmin (a furofuran type lignan), isolariciresinol (a dibenzylbutane type lignan), and cynaropicrin (a sesquiterpene lactone). The results suggest that savinin may act as an active principle in the reported biological activities of P. santalinus, such as antiinflammatory effect, by mediation of the butyrolactone ring as a valuable pharmacophore.