Active site-directed protein regulation

Regulation of protein function is vital for the control of cellular processes. Proteins are often regulated by allosteric mechanisms, in which effecters bind to regulatory sites distinct from the active sites and alter protein function. Intrasteric regulation, directed at the active site and thus the counterpart of allosteric control, is now emerging as an important regulatory mechanism.

Encapsulation of lemon oil by paste method using beta-cyclodextrin: Encapsulation efficiency and profile of oil volatiles

Microencapsulation of lemon oil was undertaken by kneading with beta-cyclodextrin, at a beta-cyclodextrin to lemon oil ratio of 88:12 (w/w). The resulting paste samples of the complex were vacuum- or spray-dried. Ten selected lemon oil flavor volatiles (alpha-pinene, sabinene, beta-pinene, beta-myrcene, limonene, gamma-terpinene, terpinolene, linalool, neral, and geranial) in the complex were analyzed periodically after 1, 2, 5, 10, 15, 20, and 30 min of kneading time. The results indicated that the levels of these volatiles were not significantly different (P > 0.05) irrespective of mixing time or type of the drying (vacuum- or spray-drying) used. An optimum mixing time was found to be 15 min, at which time the maximum encapsulation of lemon oil (97.7 mg/g of beta-cyclodextrin) was obtained in the complex powder.

Novel murine myeloid cell lines that exhibit a differentiation switch in response to IL-3 or GM-CSF, or to different constitutively active mutants of the CM-CSF receptor beta subunit

Several activating mutations have recently been described in the common beta subunit for the human interleukin(IL)-3, IL-5, and granulocyte-macrophage colony-stimulating factor (GM-CSF) receptors (h beta c), Two of these, FI Delta and 1374N, result, respectively, in a 37-amino acid duplication and an isoleucine-to-asparagine substitution in the extracellular domain. A third, V449E, leads to valine-to-glutamic acid substitution in the transmembrane domain. Previous studies have shown that when expressed in murine hemopoietic cells in vitro, the extracellular mutants can confer factor independence on only the granulocyte-macrophage lineage while the transmembrane mutant can do so to all cell types of the myeloid and erythroid compartments. To further study the signaling properties of the constitutively active hpc mutants, we have used novel murine hemopoietic cell lines, which we describe in this report. These lines, FDB1 and FDB2, proliferate in murine IL-3 and undergo granulocyte-macrophage differentiation in response to murine GM-CSF, We find that while the transmembrane mutant, V449E, confers factor-independent proliferation on these cell lines, the extracellular hpc mutants promote differentiation. Hence, in addition to their ability to confer factor independence on distinct cell types, transmembrane and extracellular activated h beta c mutants deliver distinct signals to the same cell type. Thus, the FDB cell lines, in combination with activated h beta c mutants, constitute a powerful new system to distinguish between signals that determine hemopoietic proliferation or differentiation. (C) 2000 by The American Society of Hematology.

The biologically active iron chelators 2-pyridylcarboxaldehyde isonicotinoylhydrazone, 2-pyridylcarboxaldehyde benzoylhydrazone monohydrate and 2-furaldehyde isonicotinoylhydrazone

In the crystal structures of the respective title compounds, C12H10N4O, C13H11N3O . H2O and C11K9N3O2, variations in the torsion angles of the aromatic pyridyl and benzoyl groups are observed, and the disposition of the heterocyclic aldehyde is shown to be influenced by the ring size of this group.

Synthesis of silica films at the air/water interface: Effect of template chain length and ionic strength

We have grown surfactant-templated silicate films at the air-water interface using n-alkyltrimethylammonium bromide and chloride in an acid synthesis with tetraethyl orthosilicate as the silicate source. The films have been grown with and without added salt (sodium chloride, sodium bromide) and with n-alkyl chain lengths from 12 to 18, the growth process being monitored by X-ray reflectometry. Glassy, hexagonal, and lamellar structures have been produced in ways that are predictable from the pure surfactant-water phase diagrams. The synthesis appears to proceed initially through an induction period characterized by the accumulation of silica-coated spherical micelles near the surface. All syntheses, except those involving C(12)TACl, show a sudden transformation of the spherical micellar phase to a hexagonal phase. This occurs when the gradually increasing ionic strength and/or changing ethanol concentration is sufficient to change the position of boundaries within the phase diagram. A possible mechanism for this to occur may be to induce a sphere to rod transition in the micellar structure. This transformation, as predicted from the surfactant-water phase diagram, can be induced by addition of salts and is slower for chloride than bromide counteranions. The hexagonal materials change in cell dimension as the chain length is changed in a way consistent with theoretical model predictions. All the materials have sufficiently flexible silica frameworks that phase interconversion is observed both from glassy to hexagonal and from hexagonal, to lamellar and vice versa in those surfactant systems where multiple phases are found to exist.

Functional analysis, using in vitro mutagenesis, of amino acids located in the phenylalanine hydroxylase active site

The 3-dimensionaI structure determination of rat phenylalanine hydroxylase (PAH) has identified potentially important amino acids lining the active site cleft with the majority of these having hydrophobic side-chains including several with aromatic side chains. Here we have analyzed the effect on rat PAH enzyme kinetics of in vitro mutagenesis of a number of these amino acids lining the PAH active site. Mutation of F299, Y324, F331, and Y343 caused a significant decrease in enzyme activity but no change in the K-m for substrate or cofactor. me conclude that these aromatic residues are essential for activity but are not significantly involved in binding of the substrate or cofactor. in contrast the PAH mutant, S349T, showed an 18-fold increase in K-m for phenylalanine, showing the first functional evidence that this residue was binding at or near the phenylalanine binding site. This confirms the recently published model for the binding of phenylalanine to the PAH active site that postulated S349 interacts with the amino group on the main chain of the phenylalanine molecule. This result differs with that found for the equivalent mutation (S395T), in the closely related tyrosine hydroxylase, which had no effect on substrate K-m, showing that while the architecture of the two active sites are very similar the amino acids that bind to the respective substrates are different. (C) 2000 Academic Press.

Attention Deficit Hyperactivity Disorder and substance use disorders: is there a causal link?

Attention-deficit hyperactivity disorder (ADHD), characterized by restless, inattentive and hyperactive behaviours, is a relatively common childhood disorder that affects approximately 5% of the general population. There has been controversy about whether ADHD increases risks of developing substance use disorders. The available evidence suggests that, in the absence of conduct disorder, ADHD is not associated with an increased risk of substance use problems in males. There is only limited evidence on the role of ADHD in the aetiology of substance use disorders among females. While ADHD has traditionally been considered as a childhood disorder, it may also occur in adults; research needs to examine the extent to which ADHD in adulthood increases the risk of substance use disorders.

The relationship between cannabis use and other substance use in the general population

This study examined if (1) there is an association in the general population between cannabis use, DSM-IV abuse and dependence, and other substance use and DSM-IV substance abuse/dependence; (2) if so, is it explained by demographic characteristics or levels of neuroticism? It used data from the Australian National Survey of Mental Health and Well-Being (NSMHWB), a stratified, multistage probability sample of 10641 adults, representative of the general population. DSM-IV diagnoses of substance abuse and dependence were derived using the Composite International Diagnostic Interview (CIDI). There was a strong bivariate association between involvement with cannabis use in the past 12 months and other substance use, abuse and dependence. In particular, cannabis abuse and dependence were highly associated with increased risks of other substance dependence. These associations remained after including other variables in multiple regression. Cannabis use without disorder was strongly related to other drug use, an association that was not explained by other variables considered here. The high likelihood of other substance use and substance use disorders needs to be considered among persons seeking treatment for cannabis use problems. (C) 2001 Elsevier Science Ireland Ltd. All rights reserved.

Hair morphine concentrations of fatal heroin overdose cases and living heroin users

Aims To compare heroin and other opiate use of heroin overdose fatalities, current street heroin users and drug-free therapeutic community clients. Design Hair morphine concentrations that assess heroin use and other opiate use in the 2 months preceding interview or death were compared between heroin overdose fatalities diagnosed by forensic pathologists (fOD) (n = 42), current street heroin users (CU) (n = 100) and presumably abstinent heroin users in a drug-free therapeutic community (TC) (n = 50). Setting Sydney, Australia. Findings The mean age and gender breakdown of the three samples were 32.3 years, 83% male (FOD), 28.7 years, 58% male (CU) and 28.6 years, 60% male (TC). The median blood morphine concentration among the FOD cases was 0.35 mg/l, and 82% also had other drugs detected. There were large differences between the three groups in hair morphine concentrations, with the CU group (2.10 ng/mg) having concentration approximately four times that of the FOD group (0.53 ng/mg), which in turn had a concentration approximately six times that of the TC group (0.09 ng/mg). There were no significant differences between males and females in hair concentrations within any of the groups. Hair morphine concentrations were correlated significantly with blood morphine concentrations among FOD cases (r = 0.54), and self-reported heroin use among living participants (r = 0.57). Conclusions The results indicate that fatal cases had a lower degree of chronic opiate intake than the active street users, but they were not abstinent during this period.

Patterns of co-morbidity between alcohol use and other substance use in the Australian population

The present study describes patterns of co-morbidity between alcohol use and other substance use problems in the Australian population using data from the 1997 National Survey of Mental Health and Well-Being. Multiple regression analyses examined whether the observed associations between alcohol and other drug use disorders were explained by other variables, including demographic characteristics and neuroticism. We also assessed whether the presence of co-morbid substance use disorders affected treatment seeking for a mental health problem. Alcohol use was related strongly to the use of other substances. Those who did not report alcohol use within the past 12 months were less likely to report using tobacco, cannabis, sedatives, stimulants or opiates. Higher rates again were observed among those with alcohol use disorders: half (51%) of those who were alcohol-dependent were regular tobacco smokers, one-third had used cannabis (32%); 15% reported other drug use; 15% met criteria for a cannabis use disorder and 7% met criteria for another drug use disorder. These associations were not accounted for by the demographic and other variables considered here. Co-morbid substance use disorders (sedatives, stimulants or opioids) predicted a high likelihood of seeking treatment for a mental health problem among alcohol-dependent people.