An overview of regional and local characteristics of aerosols in South Africa using satellite, ground, and modeling data.

We present a comprehensive overview of particulate air quality across the five major metropolitan areas of South Africa (Cape Town, Bloemfontein, Johannesburg and Tshwane (Gauteng Province), the Industrial Highveld Air Quality Priority Area (HVAPA), and Durban), based on a decadal (1 January 2000 to 31 December 2009) aerosol climatology from multiple satellite platforms and detailed analysis of ground-based data from 19 sites throughout Gauteng Province. Satellite analysis was based on aerosol optical depth (AOD) from MODIS Aqua and Terra (550 nm) and MISR (555 nm) platforms, Ångström Exponent (α) from MODIS Aqua (550/865 nm) and Terra (470/660 nm), ultraviolet aerosol index (UVAI) from TOMS, and results from the Goddard Ozone Chemistry Aerosol Radiation and Transport (GOCART) model. At continentally influenced sites, AOD, α, and UVAI reach maxima (0.12-0.20, 1.0-1.8, and 1.0-1.2, respectively) during austral spring (September-October), coinciding with a period of enhanced dust generation and the maximum integrated intensity of close-proximity and subtropical fires identified by MODIS Fire Information for Resource Management System (FIRMS). Minima in AOD, α, and UVAI occur during winter. Results from ground monitoring indicate that low-income township sites experience by far the worst particulate air quality in South Africa, with seasonally averaged PM10 concentrations as much as 136 % higher in townships that in industrial areas. We report poor agreement between satellite and ground aerosol measurements, with maximum surface aerosol concentrations coinciding with minima in AOD, α, and UVAI. This result suggests that remotely sensed data are not an appropriate surrogate for ground air quality in metropolitan South Africa.

Chemical composition of gas- and aerosol-phase products from the photooxidation of naphthalene.

The current work focuses on the detailed evolution of the chemical composition of both the gas- and aerosol-phase constituents produced from the OH-initiated photooxidation of naphthalene under low- and high-NO(x) conditions. Under high-NO(x) conditions ring-opening products are the primary gas-phase products, suggesting that the mechanism involves dissociation of alkoxy radicals (RO) formed through an RO(2) + NO pathway, or a bicyclic peroxy mechanism. In contrast to the high-NO(x) chemistry, ring-retaining compounds appear to dominate the low-NO(x) gas-phase products owing to the RO(2) + HO(2) pathway. We are able to chemically characterize 53-68% of the secondary organic aerosol (SOA) mass. Atomic oxygen-to-carbon (O/C), hydrogen-to-carbon (H/C), and nitrogen-to-carbon (N/C) ratios measured in bulk samples by high-resolution electrospray ionization time-of-flight mass spectrometry (HR-ESI-TOFMS) are the same as the ratios observed with online high-resolution time-of-flight aerosol mass spectrometry (HR-ToF-AMS), suggesting that the chemical compositions and oxidation levels found in the chemically-characterized fraction of the particle phase are representative of the bulk aerosol. Oligomers, organosulfates (R-OSO(3)), and other high-molecular-weight (MW) products are not observed in either the low- or high-NO(x) SOA; however, in the presence of neutral ammonium sulfate seed aerosol, an organic sulfonic acid (R-SO(3)), characterized as hydroxybenzene sulfonic acid, is observed in naphthalene SOA produced under both high- and low-NO(x) conditions. Acidic compounds and organic peroxides are found to account for a large fraction of the chemically characterized high- and low-NO(x) SOA. We propose that the major gas- and aerosol-phase products observed are generated through the formation and further reaction of 2-formylcinnamaldehyde or a bicyclic peroxy intermediate. The chemical similarity between the laboratory SOA and ambient aerosol collected from Birmingham, Alabama (AL) and Pasadena, California (CA) confirm the importance of PAH oxidation in the formation of aerosol within the urban atmosphere.

Prevalence and phenotype consequence of FRAXA and FRAXE alleles in a large, ethnically diverse, special education-needs population.

We conducted a large population-based survey of fragile X (FRAXA) syndrome in ethnically diverse metropolitan Atlanta. The eligible study population consisted of public school children, aged 7-10 years, in special education-needs (SEN) classes. The purpose of the study was to estimate the prevalence among whites and, for the first time, African Americans, among a non-clinically referred population. At present, 5 males with FRAXA syndrome (4 whites and 1 African American), among 1,979 tested males, and no females, among 872 tested females, were identified. All males with FRAXA syndrome were mentally retarded and had been diagnosed previously. The prevalence for FRAXA syndrome was estimated to be 1/3,460 (confidence interval [CI] 1/7,143-1/1,742) for the general white male population and 1/4, 048 (CI 1/16,260-1/1,244) for the general African American male population. We also compared the frequency of intermediate and premutation FRAXA alleles (41-199 repeats) and fragile XE syndrome alleles (31-199 repeats) in the SEN population with that in a control population, to determine if there was a possible phenotype consequence of such high-repeat alleles, as has been reported previously. No difference was observed between our case and control populations, and no difference was observed between populations when the probands were grouped by a rough estimate of IQ based on class placement. These results suggest that there is no phenotype consequence of larger alleles that would cause carriers to be placed in an SEN class.

Nicardipine versus nitroprusside for controlled hypotension during spinal surgery in adolescents.

Nicardipine or nitroprusside was used to induce controlled hypotension in healthy adolescents with idiopathic scoliosis undergoing spinal fusion. Twenty patients were randomly assigned to the nitroprusside (N) or nicardipine (C) group. All patients received a standardized anesthetic. A target mean arterial blood pressure (MAP) of 60 mm Hg was achieved by varying the vasoactive infusions only. Moderate hemodilution (PCV = 25) and intraoperative blood salvage were used in all cases. Hemodynamic variables, blood loss, occurrence of reflex tachycardia, and reversibility of the hypotensive state were compared between the two groups. Significant differences were observed between the two groups in the amount of blood loss and reversibility of the hypotensive state. Group C had less blood loss (761 +/- 199 mL) than Group N (1297.5 +/- 264, P < or = .05). Time to restoration of baseline MAP was longer with Group C (26.8 +/- 4.0 min) than Group N (7.3 +/- 1.1 min, P < or = 0.001). Both drugs rapidly achieved a stable, controlled hypotensive state and an acceptable operating field. There was no statistically significant difference between groups with respect to the amount of crystalloid administered or urine output. These results suggest that nicardipine is a safe, effective drug for controlled hypotension in this population and that it may offer the significant advantage of reduced blood loss in these patients.

Differential effects of GTP gamma S on acid and pepsinogen secretion by permeable gastric glands.

Gastric glands isolated from rabbit stomach were permeabilized with Staphylococcus aureus alpha-toxin. Acid secretion by parietal cells, as measured by the accumulation of weak base, was inhibited by incubation with alpha-toxin but could be restored by addition of exogenous ATP (1 mM). The permeable glands were found to retain acid secretory responses to receptor-linked secretagogues, histamine and carbachol, as well as to intracellular mediators, forskolin and 8-bromoadenosine 3',5'-cyclic monophosphate, indicating the presence of intact, functional intracellular coupling mechanisms. Both basal and stimulated acid secretion by the permeable glands were blocked by the Mg2+ chelator, trans-1,2-diaminocyclohexane-N,N,N',N'-tetraacetic acid (CDTA; 5 mM), whereas CDTA had no effect on nonpermeabilized glands. These results are interpreted to show that alpha-toxin permeabilizes parietal cells to moderate sized molecules without causing a loss of critical intracellular components. The acid secretory responses to histamine and carbachol persisted in media containing low ( < 50 nM) levels of free Ca2+ buffered by 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (0.5 mM), indicating that changes in bulk Ca2+ are not required for these responses. Inclusion of the nonhydrolyzable analogue of GTP, guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S; 100 microM), resulted in inhibition of spontaneous acid secretion, blocked responses to all agents tested, and inhibited stimulated acid secretion. GTP gamma S had no effect on nonpermeabilized glands. No effects on acid secretion by either permeable or nonpermeable glands were observed with GTP, guanosine diphosphate, or guanosine 5'-O-(2-thiodiphosphate). GTP gamma S had no effect on H+ gradient formation by gastric membrane vesicles, showing that it does not inhibit the gastric H(+)-K(+)-adenosinetriphosphatase directly. These results are interpreted to show that GTP gamma S interacts at a postreceptor site to inhibit or reverse a critical step in stimulus-secretion coupling in parietal cells. In contrast to the effect on parietal cells, GTP gamma S was found to stimulate pepsinogen secretion by alpha-toxin-permeabilized chief cells. The differential effects of GTP gamma S on acid and pepsinogen secretions suggest unique roles for GTP binding proteins in these two secretory processes. The use of alpha-toxin-permeabilized gastric glands should prove useful in defining the stimulus-secretion coupling mechanisms involved in both acid and pepsinogen secretions.

Nicardipine for controlled hypotension during spinal surgery.

Nicardipine is the first intravenously administered dihydropyridine calcium channel blocker. Its primary physiologic actions include vasodilatation with limited effects on the inotropic and dromotropic function of the myocardium. Several reports have documented its use in adult patients for pharmacologic control of blood pressure. We present our experience with nicardipine as an agent for controlled hypotension during spinal surgery in 24 children. After the induction of general anesthesia, nicardipine was started at 5 (22 patients) or 10 micrograms/kg/min (two patients). The target mean arterial pressure (MAP) of 55-65 mm Hg was reached in 5.1 +/- 2.1 min (range, 2-10). Intraoperative infusion requirements to maintain the target MAP varied from 0.5 to 7 micrograms/kg/min (mean, 2.5 +/- 1.1). No adverse effects related to nicardipine were noted. Nicardipine appears to be an effective agent for controlled hypotension in children. Future studies are required to determine its advantages/disadvantages compared with more commonly used agents such as sodium nitroprusside or adrenergic antagonists.

Analgesia after bilateral myringotomy and placement of pressure equalization tubes in children: acetaminophen versus acetaminophen with codeine.

Despite the brief nature of the procedure with limited tissue trauma, some form of analgesia is required in most children after bilateral myringotomy and placement of pressure equalization (PE) tubes. Previous studies have demonstrated the relative inefficacy of acetaminophen and nonsteroidal antiinflammatory drugs (NSAIDs), with 30%-55% of patients requiring supplemental postoperative analgesia. We undertook a prospective study evaluating the efficacy of the preoperative administration of oral acetaminophen (15 mg/kg) versus acetaminophen (10 mg/kg) and codeine (1 mg/kg). Fifty ASA grade I or II patients were randomized to receive oral midazolam premedication (0.7 mg/kg) mixed in either acetaminophen or acetaminophen with codeine elixir. Anesthesia was induced and maintained with halothane in nitrous oxide and oxygen. Postoperative pain was assessed at four times during the postoperative course using an objective pain scale. The two groups were similar with respect to age, weight, gender, duration of anesthesia, and duration of the surgical procedure. The patients who received acetaminophen with codeine had lower pain scores at all four points when compared with patients who received acetaminophen. None of the 25 patients who received acetaminophen with codeine required supplemental analgesics compared with 12 of 25 who received acetaminophen. No adverse effects were noted in either group. We conclude that the preoperative administration of acetaminophen with codeine provides superior analgesia after bilateral myringotomy and placement of PE tubes.

The pharmacology of the gastric acid pump: the H+,K+ ATPase.

The gastric H+,K+ ATPase--the gastric acid pump--is the molecular target for the class of antisecretory drugs called the proton-pump inhibitors (PPIs). These compounds--omeprazole, lansoprazole, and pantoprazole--contain, as their core structure, 2-pyridyl methylsulfinyl benzimidazole. The H+,K+ ATPase is a heterodimer composed of a 1034-amino acid catalytic alpha peptide and a glycosylated 291-amino acid beta subunit. The alpha subunit probably contains 10 membrane-spanning sequences; the beta, a single transmembrane segment. The PPIs have a pKa of about 4.0; hence they accumulate only in the acidic secretory canaliculus of the stimulated parietal cell. Here they undergo conversion to a cationic sulfenamide, which then reacts with available cysteines on the extracytoplasmic face of the alpha subunit. Omeprazole reacts and forms disulfide bonds with cys813(822) and cys892; lansoprazole, with cys813(822), cys892, and cys321; and pantoprazole, with cys813 and -822. The antisecretory effect of the drugs reflects their short plasma half-life (approximately 60 min), the number of active pumps during that time, and the recovery of pumps following biosynthesis and reversal of inhibition. These drugs also show synergism with either amoxicillin or clari- thromycin in eradicating Helicobacter pylori, an organism shown to be important in duodenal and gastric ulcer disease. Their action is probably due to elevation of pH in the environment of the organism, rather than to any direct action.

Cloning and expression of the rabbit gastric CCK-A receptor.

Cholecystokinin stimulates pancreatic zymogen secretion by binding with high affinity to a receptor on the pancreatic acinar cell. This receptor has been cloned and shown to be a CCK-A subtype. CCK also stimulates pepsinogen secretion from the gastric chief cell with high affinity. Using polymerase chain reaction with primers from the known sequence of the rat pancreatic CCK-A receptor cDNA, we prepared a 600 bp product from rat and rabbit stomach cDNA. From Southern analysis these represented a fragment of a gastric CCK-A receptor. PCR was then used to amplify a rabbit lambda ZAP II gastric epithelial cDNA library with the same primers, and the product was identified by sequencing as representing a CCK-A receptor fragment. When this PCR product was used to screen the library, ten positive clones were identified in a screening of 4.10(5) plaques, and several of these were sequenced. All had essentially the same sequence contained within 2 of these clones consisted of 427 amino acids and was 92% homologous (87% identity) to the known rat pancreatic CCK-A sequence but only 43% homologous to the gastric CCK-B sequence. The cDNA was subcloned into a pcDNA1 expression vector and transiently expressed in the human embryonic kidney cell line, HK 293. The responses of intracellular Ca2+ in these transfected cells to CCK and gastrin were monitored using video imaging. On the average 40% of the cells responded to CCK-8 by a transient elevation of [Ca2+]i followed by a steady state plateau. CCK was a high and gastrin a low affinity ligand for this signal, corresponding to the actions of these ligands on pepsinogen secretion from chief cells and somatostatin release from D cells. Hence from sequence and second messenger responses, the clone represents the CCK-A receptor presumably responsible for pepsinogen secretion by gastric chief cells and somatostatin release from gastric D cells.

Caudal epidural block for analgesia following herniorrhaphy with laparoscopy in children.

This study prospectively evaluated the efficacy of caudal epidural block in providing analgesia following inguinal herniorrhaphy and laparoscopy. Laparoscopy was used only to inspect the contralateral side to determine if a second hernia was present. No surgical manipulation was performed through the telescope. Following mask induction with halothane in nitrous oxide and oxygen, a caudal epidural block was performed with 1.2 mL/kg of 0.25% bupivacaine. Pain scores were obtained at four points during the in-hospital postoperative course, and the need for supplemental analgesic agents was assessed. A total of 45 patients were studied. Caudal epidural block could not be performed in 1 patient, and this patient was excluded from further consideration. There were 34 boys and 10 girls, ranging in age from 2 to 84 months (mean +/- SD 37.4 +/- 18.2 months) and weighing from 3.4 to 34 kg (mean +/- SD 14.2 +/- 5.8 kg). Thirty-six of 44 patients (82%) did not require supplemental analgesic agents during their in-hospital postoperative course and had pain scores of 2 or less at all four evaluation points. Six of 8 patients required a single dose of intravenous fentanyl (0.5 microgram/kg) to maintain scores of 2 or less. No significant complications related to caudal epidural block were noted in any patient. Caudal epidural block provides effective analgesia following inguinal herniorrhaphy and laparoscopy in children.

Partial agonism by gastrin for a cholecystokinin receptor mediating pepsinogen secretion.

Isolated gastric glands from rabbit were used to characterize the functional cholecystokinin (CCK)-like peptide receptors that mediate pepsinogen secretion. Pepsinogen secretion was stimulated by both CCK octapeptide sulfate (CCK-8) and A-71378, a selective CCK-A-type receptor agonist, with similar mean effective doses (1.0 and 0.8 nM, respectively). Compared with CCK-8, gastrin-17 (G-17-I) showed reduced potency and only partial efficacy for stimulation of pepsinogen secretion while inhibiting the maximal CCK-8-stimulated response. The nonpeptide inhibitors, asperlicin and L-364,718, inhibited pepsinogen secretion with identical pA2 values for antagonism of both CCK and gastrin, indicating that both peptides interact with the same functional receptor. Specific binding of [3H]CCK-8 to isolated chief cell membranes was displaced fully by both CCK and gastrin, indicating full receptor occupancy by both peptides. A novel synthetic peptide analogue, pseudogastrin [(Glu)5-Ala-Tyr-Nle-Gly-Trp-Nle-Asp-Phe-NH2], was used to investigate the structural basis for the lower potency and efficacy of G-17-I. The potency of CCK and gastrin analogues for pepsinogen secretion was found to be dependent on both sulfation of a tyrosine residue and the position of the tyrosine residue relative to the COOH-terminal phenylalanine amide. The efficacy appears to be determined partially by the extended NH2-terminal sequence of G-17-I. The results of the present study are interpreted to show that pepsinogen secretion is mediated by a CCK-A-type receptor and gastrin acts at the same receptor as a partial agonist.

Membrane topology and omeprazole labeling of the gastric H+,K(+)-adenosinetriphosphatase.

The gastric H+,K(+)-ATPase is an alpha beta heterodimer with close homology to the Na+,K(+)-ATPase. Digestion of intact cytoplasmic-side-out vesicles at a trypsin to protein ratio of 1/4 removed most of the cytoplasmic protein, leaving membrane-spanning pairs in high yield. These were visualized on gels and poly(vinylidene difluoride) (PVDF) membranes by sodium dodecyl sulfate solubilization of the membrane-embedded segments and labeling of the cysteine residues with fluorescein maleimide prior to electrophoresis. The membrane-spanning residues of the alpha subunit were found between positions 104 and 162 (M1/M2), 291 and 358(M3/M4), 776 and 835 (M5/M6), and 853 and 946 (M7/M8). Although this method did not detect membrane retention of the hydrophobic sequences subsequent to position 946, it provided biochemical evidence for at least eight membrane segments in the catalytic subunit. Intact vesicles containing this enzyme transport acid in the presence of KCl, valinomycin, and MgATP. Omeprazole accumulates in these acidified vesicles and converts to a cationic sulfenamide. This forms disulfides with accessible cysteines. The reaction with this extracytoplasmic thiol reagent inhibits ATPase activity. Full inhibition was obtained with a stoichiometry of 2.2 mol of omeprazole bound/mg of protein. Only the alpha subunit was labeled. The cysteines reacting with omeprazole were defined by proteolytic cleavage of 3H- or 14C-omeprazole-labeled enzyme followed by peptide sequencing of fragments separated on tricine gradient gels and transferred to PVDF membranes. Tryptic digestion at a 1/40 trypsin to protein ratio in the presence of ligands that stabilize the E2P form of the enzyme produced two large fragments, one of 68 kDa stretching from Glu47 to probably Arg666 that contained minor labeling and the other of 333 kDa beginning at Ala671 and extending to probably Arg946 that contained greater than 85% of the label. Digestion of labeled vesicles at 1/75 or 1/4 trypsin to protein ratios gave radioactive patterns consistent with labeling at Cys813 and/or Cys822 and at Cys892 and/or Cys927 and/or Cys938. V8 protease digestion of the solubilized alpha subunit produced a fragment extending from Ser838 to possible Asp900 that was omeprazole-labeled, showing that Cys892 was labeled and Cys927 and Cys938 were not. Hence, omeprazole labels the H+,K(+)-ATPase at cysteines within the M5/M6 and M7/M8 regions of the alpha subunit, accounting for its inhibitory action in vivo and in vitro.

The site of acid secretion in the mammalian parietal cell.

Initiation of acid secretion in the gastric mucosa is accompanied by a morphological transformation in which the acid pump, the H+/K(+)-ATPase, translocates from a cytoplasmic vesicular location to the secretory surface lining the canaliculi. Associated with the morphological changes, activation of K+ and Cl- pathways are necessary to supply K+ to the extracytoplasmic face of the pump. Although the pump in the secretory membrane is known to secrete acid, it is not known whether activation of the KCl pathway occurs in the tubulovesicular membrane prior to the formation of the canaliculus, or when the pump is in the secretory membrane. The cellular site of activation of acid secretion in the rabbit gastric parietal cell was investigated using the covalent binding of [3H]omeprazole as a probe of acid secretion in rabbit gastric glands that were undergoing stimulation in vitro. This compound depends on an acidic environment for activation and covalent binding to the H+/K(+)-ATPase. Electron microscopic autoradiography showed that activation of the enzyme occurred only when it was present in the canalicular membrane and not when it was present in the cytoplasmic tubulovesicular membrane. Hence there is likely to be a physical separation of K+ and/or Cl- pathways from the ATPase in the resting cell, and stimulation of acid secretion is dependent on colocalization of these pathways in the canalicular membrane.

Topology and sites in the H,K-ATPase.

Understanding the membrane topology of the EP-type pumps has been approached largely by analysis of hydrophobicity plots, which are confusing in the COOH-terminal third of the proteins. Each pair of predicted membrane-spanning segments with the extracytoplasmic loop contains at least one cysteine, allowing fluorescent labeling of these regions of the enzymes by cysteine reagents once the cytoplasmic domain has been removed. The membrane segment arrangement of the H,K and sr Ca ATPases was investigated by tryptic cleavage of intact cytoplasmic face-out vesicles. This was followed by fluorescein or coumarin maleimide labeling of the SDS solubilized residual membrane fragments, tricine gradient gel separation, and sequencing. The presence of four membrane-spanning pairs was demonstrated for the alpha subunit of the H,K-ATPase, with no membrane retention of H9 and H10, although H9 has four cysteines based on cDNA sequencing. A similar observation was made for the Ca pump, except that fluorescein-labeled H9 was detected in the membrane with a molecular weight of 4 kD, showing that cleavage had occurred at lys958 predicted to be extracytoplasmic in a 10 membrane segment model. It seems likely that for both these enzymes the membrane domain contains only 8 alpha helical spanning segments. Cleavage at ala236 in the beta subunit was found only in leaky, not in ion-tight vesicles, arguing for a single membrane segment in this subunit. In the H,K-ATPase additional evidence for the presence and arrangement of the first, third, and fourth pair of segments was obtained by labeling the intact enzyme with extracytoplasmic inhibitory reagents. The K competitive reagent, an imidazopyridine, MeDAZIP+, labeled the first pair of membrane segments. The acid-activated SH reagent class, the pyridinyl methyl sulfinyl benzimidazoles, labeled cysteines 813 and 822 in the M5/M6 region as well as cysteine 892 in the extracytoplasmic loop between M7 and M8. No labeling of the beta subunit was found, indicating the presence of three disulfide bonds in the extracytoplasmic domain of this subunit. Both sets of extracytoplasmic reagents are predicted to bind close to the fatty acid/phospholipid head group interface. Inhibition by these reagents shows that conformational changes are transmitted between cytoplasmic and extracytoplasmic domains.

Acid secretion and the H,K ATPase of stomach.

The regulation of acid secretion was clarified by the development of H2-receptor antagonists in the 1970s. It appears that gastrin and acetylcholine exert their effects on acid secretion mainly by stimulation of histamine release from the enterochromaffin-like (ECL) cell of the fundic gastric mucosa. The isolated ECL cell of rat gastric mucosa responds to gastrin/cholecystokinin (CCK), acetylcholine, and epinephrine with histamine release and to somatostatin and R-alpha-methyl histamine by inhibition of histamine release. Histamine and acetylcholine stimulate the parietal cell by elevation of cAMP or [Ca]i by activation of H2 or M3 receptors, respectively. These independent pathways converge to activate the gastric acid pump, the H+,K+ ATPase. Activation is a function of the association of the ATPase with a potassium chloride transport pathway that occurs in the membrane of the secretory canaliculus of the parietal cell. Hence the secretory canaliculus is the site of acid secretion, the acid being pumped into the lumen of the canaliculus. The pump is composed of two subunits, a large catalytic and a smaller glycosylated protein. This final step of acid secretion has become the target of drugs also designed to inhibit acid secretion. The target domain of the benzimidazole class of acid pump inhibitors is the extracytoplasmic domain of the pump that is secreting acid, and the target amino acids are the cysteines present in this domain. The secondary structure of the pump can be analyzed by determining trypsin-sensitive bonds in intact, cytoplasmic-side-out vesicles of the ATPase, and it has been shown that the alpha subunit has at least eight membrane-spanning segments. Omeprazole, the first acid pump inhibitor, forms a disulfide bond with cysteines in the extracytoplasmic loop between the fifth and sixth membrane-spanning segment and to a cysteine in the extracytoplasmic loop between the seventh and eight segments, preventing phosphorylation of the pump by ATP. As a result of the effective and long-lasting inhibition of acid secretion by the acid pump inhibitor, superior clinical results have been found in all forms of acid-related disease.

Characterization of the adherence of Porphyromonas gingivalis to oral streptococci.

Adherence of Porphyromonas gingivalis to strains of Streptococcus sanguis and Streptococcus mitis deposited on nitrocellulose paper was investigated. A variety of laboratory strains and clinical isolates of P. gingivalis bound to both S. sanguis and S. mitis. Binding of P. gingivalis to all but one of the streptococci was not inhibited by salivary molecules. Pretreatment of P. gingivalis with periodate and pretreatment of S. sanguis and S. mitis with pronase decreased binding, suggesting that adherence may be mediated by a protein on the streptococci interacting with a carbohydrate on P. gingivalis. Binding was not inhibited by a selection of simple sugars. The ability to adhere to early plaque bacteria such as S. sanguis and S. mitis may be important in the colonization of the mouth by P. gingivalis.

Structural aspects of the gastric H,K-ATPase.

The gastric H,K-ATPase is an alpha, beta heterodimer. The large catalytic subunit is composed, in the case of the hog enzyme, of 1033 amino acids, whereas the beta subunit is composed of about 291 amino acids and is heavily glycosylated. The membrane topology of the alpha subunit is difficult to predict using hydropathy analysis. Tryptic hydrolysis of intact, inside out vesicles followed by cysteine labelling with fluorescein-5-maleimide provided experimental evidence for an 8 membrane spanning model for the alpha subunit, between residues 104 and 162 (M1/M2), 291 and 358 (M3/M4), 776 and 835 (M5/M6), and 853 and 946 (M7/M8). No evidence was found for a pair of segments (M9/M10) towards the C terminal end of the molecule, contrary to predictions for the Na,K- and Ca-ATPases. Iodination of intact vesicles followed by carboxypeptidase Y cleavage of the C terminal tyrosines showed that the C terminal end of the alpha subunit was cytoplasmic. The epitope for antibody 146 was extracytoplasmic and located between residues 871 to 874 between M7/M8. The binding site of the K competitive imidazo-pyridine, SCH28080, was to the extracytoplasmic loop between M1 and M2, whereas the binding of the covalent SH reagent generated from acid activation of omeprazole in acid transporting vesicles was to 2 cysteines at positions 813 (or 822) and 892 predicted to be in the extracytoplasmic loops connecting M5/M6 and M7/M8, respectively. The beta subunit was only hydrolysed in broken vesicles. A fragment beginning at position 236 was liberated under these conditions only in the presence of reducing agents, showing that cysteine 210 and 263 were disulfide linked.(ABSTRACT TRUNCATED AT 250 WORDS)

Chemomechanical coupling in the gastric H,K ATPase.

The gastric H,K ATPase is investigated in terms of its secondary structure by analysis of the binding sites of the extracytoplasmic inhibitors and by tryptic cleavage of intact, inside-out gastric vesicles. The inhibitors affect phosphorylation and other partial reactions of the ATPase that depend on cytoplasmic conformational changes. The K competitive imidazopyridine, SCH28080 binds to the first pair of transmembrane segments, M1/M2, probably at phe124 and asp136. Omeprazole which generates a cationic sulfenamide in acid spaces binds to either cys813 or cys822 at one site and cys892 at the other. These cysteines are located at the membrane spanning pairs, M5/M6 and at M7/M8. Tryptic cleavage of intact inside out vesicles followed by labelling with fluorescein-5-maleimide provides direct evidence for 8 membrane spanning segments between positions 104/162 (M1/M2), 291/358(M3/M4), 776/835 (M5/M6),853/946 (M7/M8). Evidence is lacking so far for M9/M10, postulated on the basis of hydrophobicity for the Ca ATPase. Conformational studies suggest that there is interaction between the cytoplasmic loop between M4 and M5 (ATP domain) and the extracytoplasmic domain of the enzyme at the inhibitor binding sites.