Effect of manganese on alkaline phosphatase production in Bacillus sp. RK11

Summary Bacillus sp. RKll, an alkalophilic isolate from soil, produces an extracellular alkaline phosphatase. In the absence of Mn²⁺ in a complex medium, no alkaline phosphatase production or sporulation by the organism was detected. No other divalent metal could be substituted for Mn²⁺ in enzyme production or in sporulation. Manganous sulphate (70 mol) gave highest enzyme production although spore numbers continued to increase with manganous concentrations above this level. Maximum alkaline phosphatase production occurred when the metal was present at the time of inoculation but maximum spore numbers were detected when the metal was added 8–12h after inoculation. Inorganic pyrophosphatase was not associated with extracellular alkaline phosphatase, but it was detected intracellularly. Ninety-five percent of the alkaline phosphatase was detected extracellularly.     

A novel maltohexaose-forming α-amylase from Bacillus caldovelox: patterns and mechanisms of action

Summary The acidophilic, thermostable -amylase of Bacillus caldovelox displays a unique end-product profile and action pattern on starch. Maltohexaose is preferentially produced, with a maximum yield of 40–44% (w/w) from 35% (w/v) starch and dextrins (DE 9 and DE 18). Maltohexaose, the initial product of 1% (w/v) starch and 35% (w/v) dextrin (DE 42) hydrolysis, is subsequently converted into maltopentaose with a maximum yield of 30% (w/w). This reaction does not involve glucose production. Substrates were hydrolysed from the non-reducing end by either a uni- or multimolecular mechanism, with no hydrolysis of maltohexaose or smaller sugars. The K _m values for soluble starch and maltoheptaose were 4.68 mg/ml and 2.13 × 10^–2 M, respectively. Offprint requests to: C. T. Kelly     

Studies on the thermostability of the α-amylase of Bacillus caldovelox

Summary Molecular mechanisms of thermoinactivation of the thermostable -amylase of Bacillus caldovelox were examined. Monomolecular conformational processes were found to be the major causes of thermoinactivation at both pH 4.5 and 8.0. The enzyme possessed considerable additional thermostability at pH 8.0, with half-lives of 0.75 and 7.0 min at 90° C and pH 4.5 and 8.0, respectively. The amino acid composition was examined with respect to the underlying thermostability exhibited by this enzyme. The inherent thermostability exhibited may be due to the high proline content (4.47 mol%), but more likely due to the high content of residues forming hydrophobic bonds (60.89 mol%) allied to a low content of residues responsible for ionic interactions (28.34 mol%). Offprint requests to: C. T. Kelly     

Extracellular xylanolytic enzymes ofPaecilomyces varioti

Summary Paecilomyces varioti produced an extracellular xylanase and B-xylosidase when cultured in a medium containing xylan and corn steep liquor. Xylose (2%, w/v) totally inhibited production of both enzymes. The enzymes were purified and both had a pH optimum of 4.0. The xylanase had a molecular weight of 20,000, an isoelectric point of 5.2 and was inactive on all substrates tested except xylan. The -xylosidase, a glycoprotein, had a molecular weight of 67,000, an isoelectric point of 4.0 and had highest activity on p-nitrophenyl--D-xyloside. The xylanase had a Km of 49.5 mg/ml for xylan and the -xylosidase had a Km of 5.4 mM for p-nitrophenyl--D-xyloside.     

The isomaltulose synthesising enzyme ofSerratia plymuthica

Summary An intracellular enzyme was located inSerratia plymuthica which produced isomaltulose from sucrose. The enzyme was purified giving a preparation with a specific activity of 1,285. It has pH and temperature optima of 6.0 and 30°C, respectively. The enzyme was stable retaining 100% activity after 2 weeks at 30°C. It had an isoelectric point at pH 9.0, a Mr of 79,500 and the Km for sucrose was 65.3mM. The enzyme converted 40% (w/v) sucrose to isomaltulose with an efficiency of 87%.     

A close association of RyRs with highly dense clusters of Ca2+-activated Cl- channels underlies the activation of STICs by Ca2+ sparks in mouse airway smooth muscle

Ca(2+) sparks are highly localized, transient releases of Ca(2+) from sarcoplasmic reticulum through ryanodine receptors (RyRs). In smooth muscle, Ca(2+) sparks trigger spontaneous transient outward currents (STOCs) by opening nearby clusters of large-conductance Ca(2+)-activated K(+) channels, and also gate Ca(2+)-activated Cl(-) (Cl((Ca))) channels to induce spontaneous transient inward currents (STICs). While the molecular mechanisms underlying the activation of STOCs by Ca(2+) sparks is well understood, little information is available on how Ca(2+) sparks activate STICs. In the present study, we investigated the spatial organization of RyRs and Cl((Ca)) channels in spark sites in airway myocytes from mouse. Ca(2+) sparks and STICs were simultaneously recorded, respectively, with high-speed, widefield digital microscopy and whole-cell patch-clamp. An image-based approach was applied to measure the Ca(2+) current underlying a Ca(2+) spark (I(Ca(spark))), with an appropriate correction for endogenous fixed Ca(2+) buffer, which was characterized by flash photolysis of NPEGTA. We found that I(Ca(spark)) rises to a peak in 9 ms and decays with a single exponential with a time constant of 12 ms, suggesting that Ca(2+) sparks result from the nonsimultaneous opening and closure of multiple RyRs. The onset of the STIC lags the onset of the I(Ca(spark)) by less than 3 ms, and its rising phase matches the duration of the I(Ca(spark)). We further determined that Cl((Ca)) channels on average are exposed to a [Ca(2+)] of 2.4 microM or greater during Ca(2+) sparks. The area of the plasma membrane reaching this level is <600 nm in radius, as revealed by the spatiotemporal profile of [Ca(2+)] produced by a reaction-diffusion simulation with measured I(Ca(spark)). Finally we estimated that the number of Cl((Ca)) channels localized in Ca(2+) spark sites could account for all the Cl((Ca)) channels in the entire cell. Taken together these results lead us to propose a model in which RyRs and Cl((Ca)) channels in Ca(2+) spark sites localize near to each other, and, moreover, Cl((Ca)) channels concentrate in an area with a radius of approximately 600 nm, where their density reaches as high as 300 channels/microm(2). This model reveals that Cl((Ca)) channels are tightly controlled by Ca(2+) sparks via local Ca(2+) signaling.     

Antibodies to mutated citrullinated vimentin and disease activity score in early arthritis: a cohort study

Introduction  

The aim of our study was to investigate the association between arthritic disease activity and antibodies to mutated citrullinated vimentin (anti-MCV), because such a relation has been suggested.     

In vitro zygotic embryo culture of wild Musa acuminata ssp.malaccensis and factors affecting germination and seedling growth

In vitro zygotic embryo culture of wild banana significantly increased the germination compared to greenhouse grown seeds. Embryo orientation and BAP concentration significantly affected germination rate. These factors together with gelling agent, dark and light conditions and coconut water, also showed variable effects on the number of roots per plant, root length, shoot length, number of days to root emergence and number of days to shoot emergence.