scholarly journals Purification and properties of the P2 primary alkylsulphohydrolase of the detergent-degrading bacterium pseudomonas C12B

1980 ◽  
Vol 185 (1) ◽  
pp. 23-31 ◽  
Author(s):  
J M Cloves ◽  
K S Dodgson ◽  
G F White ◽  
J W Fitzgerald
Keyword(s):  
Free Energy ◽  
Chain Length ◽  
Alkyl Chain ◽  
Competitive Inhibition ◽  
Deae Cellulose ◽  
Intact Protein ◽  
Alkyl Sulphate ◽  
Degrading Bacterium ◽  
Purification And Properties ◽  
Considerable Length

The P2 primary alkylsulphohydrolase of the soil bacterium Pseudomonas C12B was purified to homogeneity (200-250-fold) by column chromatography on DEAE-cellulose, Sephadex G-100 and butyl-agarose. The intact protein is a dimer with a mol. wt. of 160 000. Activity towards primary alkyl sulphate esters was maximal at pH 8.3, varied little in the range pH 7.8-8.7, but decreased sharply at higher pH. For a homologous series of primary alkyl sulphate substrates (C6-C12), logKm decreased linearly with increasing chain length, corresponding to a contribution to the free energy of association between enzyme and substrate of ‒2.5kJ/mol for each additional CH2 group in the alkyl chain. logKi for the competitive inhibition by secondary alkyl 2-sulphate esters followed a similar pattern (-2.4kJ/mol for each additional CH2 group) except that only n-1 carbon atoms effectively participate in hydrophobic bonding, implying that the C-1 methyl group is not involved. logKi values for inhibition primary alkanesulphonates also depended linearly on chain length but with a diminished gradient, indicating a free-energy increment of ‒1.2kJ/mol per additional CH2 group. The collective results showed the presence of a hydrophobic site on the enzyme capable of accomodating an alkyl chain of considerable length. Cationic structures (in the form of arginine, lysine or histidine), whose presence might be expected for binding the anionic sulphate group, were not detectable at the active site.

scholarly journals Further studies on the substrate specificity and inhibition of the stereospecific CS2 secondary alkylsulphohydrolase of Comamonas terrigena

1980 ◽  
Vol 191 (2) ◽  
pp. 467-473 ◽  
Author(s):  
Carol H. Barrett ◽  
Kenneth S. Dodgson ◽  
Graham F. White
Keyword(s):  
Free Energy ◽  
Chain Length ◽  
Alkyl Chain ◽  
Hydrophobic Interactions ◽  
Standard Free Energy ◽  
Alkyl Chain Length ◽  
Alkyl Sulphate ◽  
Competitive Inhibitors ◽  
Free Energy Of Binding ◽  
Comamonas Terrigena

A series of d-alkan-2-yl sulphate esters (C7–C14) were prepared by sulphation of the resolved parent alcohols by a method that entails complete retention of configuration. These sulphate esters were tested as substrates for the stereospecific CS2 secondary alkylsulphohydrolase of Comamonas terrigena. Vmax. reached a maximum with the C9 compound, whereas logKm decreased linearly as the alkyl-chain length was increased from C7 to C14. A parallel series of l-alkan-2-yl sulphates was also prepared, and these esters, together with homologous series of primary alkyl sulphates and primary alkanesulphonates, were shown to be competitive inhibitors of the CS2 enzyme. For each series of compounds, logKi values decreased linearly with increasing alkyl-chain length. Plots of chain length against the standard free energy of binding (ΔG0) of substrate and inhibitors to the CS2 enzyme showed that the standard free energy of association of a –CH2– group with the enzyme was 2.0–2.4kJ/mol for all classes of compound studied, indicating an important contribution from hydrophobic interactions to the overall binding. Plots for d-alkan-2-yl sulphate substrates and primary alkyl sulphate inhibitors were nearly coincident, suggesting that the overall interaction between a primary ester and the enzyme is the same as that between the isomeric secondary substrate and the enzyme. Plots for l-alkan-2-yl sulphate and alkanesulphonate inhibitors were very similar to each other, but were displaced by 1.5–3.0kJ/mol from that for substrate binding. This indicates that the binding of any one of these particular inhibitors involves one carbon atom fewer than the number involved in binding a substrate of the same chain length. These observations are discussed in terms of a three-point attachment of substrate to the enzyme involving the alkyl chain, sulphate group and the C-1 methyl group.

scholarly journals Primary alkylsulphatase activities of the detergent-degrading bacterium Pseudomonas C12B. Purification and properties of the P1 enzyme

1986 ◽  
Vol 236 (2) ◽  
pp. 401-408 ◽  
Author(s):  
T J Bateman ◽  
K S Dodgson ◽  
G F White
Keyword(s):  
Chain Length ◽  
Hydrophobic Interactions ◽  
Deae Cellulose ◽  
Carbon Chain ◽  
Maximum Activity ◽  
Carbon Chain Length ◽  
Streptomycin Sulphate ◽  
Degrading Bacterium ◽  
Purification And Properties ◽  
Hydrophobic Binding

The P1 primary alkylsulphatase of Pseudomonas C12B was purified 1500-fold to homogeneity by a combination of streptomycin sulphate precipitation of nucleic acids, (NH4)2SO4 fractionation and chromatography on columns of DEAE-cellulose, Sephacryl S-300 and butyl-agarose. The protein was tetrameric with an Mr of 181000-193000, and exhibited maximum activity at pH 6.1. Primary alkyl sulphates of carbon-chain length C1-C5 or above C14 were not substrates, but the intermediate homologues were shown to be substrates, either by direct assay (C6-C9 and C12) or by gel zymography (C10, C11, C13 and C14). Increasing the chain length from C6 to C12 led to diminishing Km. Values of delta G0′ for binding substrates to enzyme were dependent linearly on chain length, indicating high dependence on hydrophobic interactions. Vmax./Km values increased with increasing chain length. Inhibition by alk-2-yl sulphates and alkane-sulphonates was competitive and showed a similar dependence on hydrophobic binding. The P1 enzyme was active towards several aryl sulphates, including o-, m- and p-chlorophenyl sulphates, 2,4-dichlorophenyl sulphate, o-, m- and p-methoxyphenyl sulphates, m- and p-hydroxyphenyl sulphates and p-nitrophenyl sulphate, but excluding bis-(p-nitrophenyl) sulphate and the O-sulphate esters of tyrosine, nitrocatechol and phenol. The arylsulphatase activity was weak compared with alkylsulphatase activity, and it was distinguishable from the de-repressible arylsulphatase activity of Pseudomonas C12B reported previously. Comparison of the P1 enzyme with the inducible P2 alkylsulphatase of this organism, and with the Crag herbicide sulphatase of Pseudomonas putida, showed that, although there are certain similarities between any two of the three enzymes, very few properties are common to all three.

scholarly journals Purification and properties of the S1 secondary alkylsulphohydrolase of the detergent-degrading micro-organism, Pseudomonas C12B

1978 ◽  
Vol 169 (3) ◽  
pp. 659-667 ◽  
Author(s):  
Barbara Bartholomew ◽  
Kenneth S. Dodgson ◽  
Stephen D. Gorham
Keyword(s):  
Chain Length ◽  
Alkyl Chain ◽  
Competitive Inhibitor ◽  
Sulphate Group ◽  
Experimental Conditions ◽  
Alkyl Sulphate ◽  
Purification And Properties ◽  
Crude Preparation ◽  
A Chain ◽  
Comamonas Terrigena

The S1 secondary alkylsulphohydrolase of the detergent-degrading micro-organism, Pseudomonas C12B, was separated from other alkylsulphohydrolases and purified to homogeneity. Under the experimental conditions used the enzyme completely hydrolysed d-octan-2-yl sulphate (d-1-methylheptyl sulphate), but showed no activity towards the corresponding l-isomer. Additional evidence has been obtained to indicate that it is probably optically stereospecific for d-secondary alkyl sulphate esters with the ester sulphate group at C-2 and with a chain length of at least seven carbon atoms. Enzyme activity towards racemic samples of heptan-2-yl sulphate (1-methylhexyl sulphate), octan-2-yl sulphate and decan-2-yl sulphate (1-methylnonyl sulphate) increased with increasing chain length. l-Octan-2-yl sulphate is a competitive inhibitor of the enzyme, as are certain primary alkyl sulphates and primary alkanesulphonates. Inhibition by each of the last two types of compounds is characteristic of the behaviour of an homologous series. Inhibition increases with increasing chain length and plots of log Ki values against the number of carbon atoms in each alkyl chain show the expected linear relationship. A crude preparation of the S2 secondary alkylsulphohydrolase was used to show that this particular enzyme hydrolyses l-octan-2-yl sulphate, but is probably inactive towards the corresponding d-isomer. The similarity of the S1 and S2 enzymes to the CS2 and CS1 enzymes respectively of Comamonas terrigena was established, and some comments have been made on the possible roles of these and other alkylsulphohydrolases in the biodegradation of detergents.

Kinetic Behaviour of Amylase According to pH: A New Perspective for Starch Hydrolysis Process

2020 ◽  
Vol 16 (2) ◽  
pp. 135-144
Author(s):  
Ravneet K. Grewal ◽  
Baldeep Kaur ◽  
Gagandeep Kaur
Keyword(s):  
Metal Ions ◽  
Competitive Inhibition ◽  
Deae Cellulose ◽  
Kinetic Studies ◽  
Cost Effective ◽  
Starch Hydrolysis ◽  
Divalent Metal Ions ◽  
Activity Data ◽  
Time Saving ◽  
Alkaline Conditions

Background: Amylases are the most widely used biocatalysts in starch saccharification and detergent industries. However, commercially available amylases have few limitations viz. limited activity at low or high pH and Ca2+ dependency. Objective: The quest for exploiting amylase for diverse applications to improve the industrial processes in terms of efficiency and feasibility led us to investigate the kinetics of amylase in the presence of metal ions as a function of pH. Methods: The crude extract from soil fungal isolate cultures is subjected to salt precipitation, dialysis and DEAE cellulose chromatography followed by amylase extraction and is incubated with divalent metal ions (i.e., Ca2+, Fe2+, Cu2+, and Hg2+); Michaelis-Menton constant (Km), and maximum reaction velocity (Vmax) are calculated by plotting the activity data obtained in the absence and presence of ions, as a function of substrate concentration in Lineweaver-Burk Plot. Results: Kinetic studies reveal that amylase is inhibited un-competitively at 5mM Cu2+ at pH 4.5 and 7.5, but non-competitively at pH 9.5. Non-competitive inhibition of amylase catalyzed starch hydrolysis is observed with 5mM Hg2+ at pH 9.5, which changes to mixed inhibition at pH 4.5 and 7.5. At pH 4.5, Ca2+ induces K- and V-type activation of amylase catalyzed starch hydrolysis; however, the enzyme has V-type activation at 7mM Ca2+ under alkaline conditions. Also, K- and V-type of activation of amylase is observed in the presence of 7mM Fe2+ at pH 4.5 and 9.5. Conclusion: These findings suggest that divalent ions modulation of amylase is pH dependent. Furthermore, a time-saving and cost-effective solution is proposed to overcome the challenges of the existing methodology of starch hydrolysis in starch and detergent industries.

scholarly journals Silver Nanoparticles Stabilized with Phosphorus-Containing Heterocyclic Surfactants: Synthesis, Physico-Chemical Properties, and Biological Activity Determination

Nanomaterials ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 1883
Author(s):  
Martin Pisárčik ◽  
Miloš Lukáč ◽  
Josef Jampílek ◽  
František Bilka ◽  
Andrea Bilková ◽  
...  
Keyword(s):  
Silver Nanoparticles ◽  
Biological Activity ◽  
Zeta Potential ◽  
Chain Length ◽  
Alkyl Chain ◽  
Chemical Properties ◽  
Alkyl Chain Length ◽  
Microbial Pathogens ◽  
Physico Chemical ◽  
Phosphorus Containing

Phosphorus-containing heterocyclic cationic surfactants alkyldimethylphenylphospholium bromides with the alkyl chain length 14 to 18 carbon atoms were used for the stabilization of silver nanodispersions. Zeta potential of silver nanodispersions ranges from +35 to +70 mV, which indicates the formation of stable silver nanoparticles (AgNPs). Long-chain heptadecyl and octadecyl homologs of the surfactants series provided the most intensive stabilizing effect to AgNPs, resulting in high positive zeta potential values and smaller diameter of AgNPs in the range 50–60 nm. A comparison with non-heterocyclic alkyltrimethylphosphonium surfactants of the same alkyl chain length showed better stability and more positive zeta potential values for silver nanodispersions stabilized with heterocyclic phospholium surfactants. Investigations of biological activity of phospholium-capped AgNPs are represented by the studies of antimicrobial activity and cytotoxicity. While cytotoxicity results revealed an increased level of HepG2 cell growth inhibition as compared with the cytotoxicity level of silver-free surfactant solutions, no enhanced antimicrobial action of phospholium-capped AgNPs against microbial pathogens was observed. The comparison of cytotoxicity of AgNPs stabilized with various non-heterocyclic ammonium and phosphonium surfactants shows that AgNPs capped with heterocyclic alkyldimethylphenylphospholium and non-heterocyclic triphenyl-substituted phosphonium surfactants have the highest cytotoxicity among silver nanodispersions stabilized by the series of ammonium and phosphonium surfactants.

scholarly journals Physico-chemical and gel properties of heat-induced pasteurized liquid egg white gel: effect of alkyl chain length of alcohol

2021 ◽  
Vol 24 (1) ◽  
pp. 1229-1243
Author(s):  
Danai Charoensuk ◽  
Robert G. Brannan ◽  
Wilailuk Chaiyasit ◽  
Wanlop Chanasattru
Keyword(s):  
Chain Length ◽  
Alkyl Chain ◽  
Egg White ◽  
Alkyl Chain Length ◽  
Gel Properties ◽  
Gel Effect ◽  
Physico Chemical
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