Tryptophan 500 and Arginine 707 Define Product and Substrate Active Site Binding in Soybean Lipoxygenase-1†

Biochemistry ◽  
2004 ◽  
Vol 43 (41) ◽  
pp. 13063-13071 ◽  
Author(s):  
Viola C. Ruddat ◽  
Rakesh Mogul ◽  
Ilya Chorny ◽  
Cameron Chen ◽  
Noah Perrin ◽  
...  
Keyword(s):  
Active Site ◽  
Soybean Lipoxygenase ◽  
Site Binding

scholarly journals Molecular mechanism of uncompetitive inhibition of human placental and germ-cell alkaline phosphatase

1992 ◽  
Vol 286 (1) ◽  
pp. 23-30 ◽  
Author(s):  
M F Hoylaerts ◽  
T Manes ◽  
J L Millán
Keyword(s):  
Germ Cell ◽  
Active Site ◽  
Molecular Model ◽  
Inhibition Mechanism ◽  
Side Group ◽  
Alkaline Phosphatases ◽  
Uncompetitive Inhibition ◽  
Guanidinium Group ◽  
Hydrolysis Of ◽  
Site Binding

Placental (PLAP) and germ-cell (GCAP) alkaline phosphatases are inhibited uncompetitively by L-Leu and L-Phe. Whereas L-Phe inhibits PLAP and GCAP to the same extent, L-Leu inhibits GCAP 17-fold more strongly than it does PLAP. This difference has been attributed [Hummer & Millán (1991) Biochem. J 274, 91-95] to a Glu----Gly substitution at position 429 in GCAP. The D-Phe and D-Leu enantiomorphs are also inhibitory through an uncompetitive mechanism but with greatly decreased efficiencies. Replacement of the active-site residue Arg-166 by Ala-166 changes the inhibition mechanism of the resulting PLAP mutant to a more complex mixed-type inhibition, with decreased affinities for L-Leu and L-Phe. The uncompetitive mechanism is restored on the simultaneous introduction of Gly-429 in the Ala-166 mutant, but the inhibitions of [Ala166,Gly429]PLAP and even [Lys166,Gly429]PLAP by L-Leu and L-Phe are considerably decreased compared with that of [Gly429]PLAP. These findings point to the importance of Arg-166 during inhibition. Active-site binding of L-Leu requires the presence of covalently bound phosphate in the active-site pocket, and the inhibition of PLAP by L-Leu is pH-sensitive, gradually disappearing when the pH is decreased from 10.5 to 7.5. Our data are compatible with the following molecular model for the uncompetitive inhibition of PLAP and GCAP by L-Phe and L-Leu: after binding of a phosphorylated substrate to the active site, the guanidinium group of Arg-166 (normally involved in positioning phosphate) is redirected to the carboxy group of L-Leu (or L-Phe), thus stabilizing the inhibitor in the active site. Therefore leucinamide and leucinol are weaker inhibitors of [Gly429]PLAP than is L-Leu. During this Arg-166-regulated event, the amino acid side group is positioned in the loop containing Glu-429 or Gly-429, leading to further stabilization. Replacement of Glu-429 by Gly-429 eliminates steric constraints experienced by the bulky L-Leu side group during its positioning and also increases the active-site accessibility for the inhibitor, providing the basis for the 17-fold difference in inhibition efficiency between PLAP and GCAP. Finally, the inhibitor's unprotonated amino group co-ordinates with the active-site Zn2+ ion 1, interfering with the hydrolysis of the phosphoenzyme intermediate, a phenomenon that determines the uncompetitive nature of the inhibition.

A Comparison of Active Site Binding of 4-Quinolones and Novel Flavone Gyrase Inhibitors to DNA Gyrase

1995 ◽  
pp. 59-69 ◽  
Author(s):  
J. J. Hilliard ◽  
H. M. Krause ◽  
J. I. Bernstein ◽  
J. A. Fernandez ◽  
V. Nguyen ◽  
...  
Keyword(s):  
Active Site ◽  
Dna Gyrase ◽  
Gyrase Inhibitors ◽  
Site Binding

scholarly journals Regulation of the acto·myosin subfragment 1 interaction by troponin/tropomyosin. Evidence for control of a specific isomerization between two acto·myosin subfragment 1 states

1991 ◽  
Vol 279 (3) ◽  
pp. 711-718 ◽  
Author(s):  
D F A McKillop ◽  
M A Geeves
Keyword(s):  
Skeletal Muscle ◽  
Active Site ◽  
Actin Filaments ◽  
Thin Filaments ◽  
Closed State ◽  
Myosin Subfragment ◽  
Actomyosin Interaction ◽  
Binding Isotherms ◽  
Myosin Subfragment 1 ◽  
Site Binding

The co-operative binding of myosin subfragment 1 (S1) to reconstituted skeletal-muscle thin filaments has been examined by monitoring the fluorescence of a pyrene probe on Cys-374 of actin. The degree of co-operativity differs when phosphate, sulphate or ADP are bound to the S1 active site. Binding isotherms have been analysed according to the Geeves & Halsall [(1987) Biophys. J. 52, 215-220] model, which proposed that troponin and tropomyosin effected regulation of the actomyosin interaction by controlling an isomerization of the actomyosin complex. The data support the proposal that seven actin monomers associated with a single tropomyosin molecule act as a co-operative unit that can be in one of two states. In the ‘closed’ state myosin can bind to actin, but the subsequent isomerization is prevented. The isomerization is only allowed after the seven-actin unit is in the ‘open’ form. Ca2+ controls the proportion of actin filaments in the ‘closed’ and ‘open’ forms in the absence of myosin heads. The ratio of ‘closed’ to ‘open’ forms is approx. 50:1 in the absence of Ca2+ and 5:1 in its presence.

Active site binding modes of curcumin in HIV-1 protease and integrase

2005 ◽  
Vol 15 (14) ◽  
pp. 3364-3368 ◽  
Author(s):  
Opa Vajragupta ◽  
Preecha Boonchoong ◽  
Garrett M. Morris ◽  
Arthur J. Olson
Keyword(s):  
Active Site ◽  
Binding Modes ◽  
Hiv 1 ◽  
Site Binding

Out of the active site binding pocket for carbonic anhydrase inhibitors

2015 ◽  
Vol 51 (2) ◽  
pp. 302-305 ◽  
Author(s):  
Katia D'Ambrosio ◽  
Simone Carradori ◽  
Simona M. Monti ◽  
Martina Buonanno ◽  
Daniela Secci ◽  
...  
Keyword(s):  
Carbonic Anhydrase ◽  
Active Site ◽  
Binding Pocket ◽  
Carbonic Anhydrase Ii ◽  
Carbonic Anhydrase Inhibitors ◽  
Human Carbonic Anhydrase ◽  
Site Binding

2-Benzylsulfinylbenzoic acid binds to human carbonic anhydrase II in a mode completely different from any other class of carbonic anhydrase inhibitors investigated so far.

scholarly journals Allosteric communication in mammalian muscle aldolase

1997 ◽  
Vol 327 (3) ◽  
pp. 717-720 ◽  
Author(s):  
Jurgen SYGUSCH ◽  
Danielle BEAUDRY
Keyword(s):  
Active Site ◽  
Conformational Transition ◽  
Exchange Chromatography ◽  
Dihydroxyacetone Phosphate ◽  
Allosteric Communication ◽  
Modified Enzyme ◽  
Substrate Protection ◽  
Fructose 1,6 Bisphosphate ◽  
Single Subunit ◽  
Site Binding

Mixed disulphide formation in the presence of oxidized glutathione reversibly inactivates rabbit skeletal muscle aldolase. Inactivation is allosteric, preferentially modifying Cys-72 on the surface of the aldolase homotetramer distant from active-site locations and subunit interfaces. Ion-exchange chromatography fractionates partly inactivated aldolase into three distinct enzymic species: unmodified enzyme, inactive fully modified enzyme corresponding to one thiol reacted per subunit, and inactive singly modified enzyme in which only one thiol has reacted. Acid-precipitable enzymic intermediates formed in the presence of substrate, D-fructose 1,6-bisphosphate, and product, dihydroxyacetone phosphate, indicates that active site binding is unaffected upon modification. The absence of enamine carbanion formation in the presence of substrate but not product is consistent with mixed disulphide formation's blocking -C-C- cleavage and/or subsequent D-glyceraldehyde 3-phosphate release. Inactivation upon single subunit modification and substrate protection against modification denotes that the blocked step is associated with a long-range conformational transition involving highly co-operative subunit behaviour.

Lipoxygenase Inhibitory Activity of 6-Pentadecanylsalicylic Acid without Prooxidant Effect

2010 ◽  
Vol 5 (1) ◽  
pp. 1934578X1000500 ◽  
Author(s):  
Isao Kubo ◽  
Tae Joung Ha ◽  
Kuniyoshi Shimizu
Keyword(s):  
Active Site ◽  
Competitive Inhibitor ◽  
Anacardic Acid ◽  
Soybean Lipoxygenase ◽  
Inhibition Kinetics ◽  
Iron Ion ◽  
Prooxidant Effect ◽  
Specific Structural Feature ◽  
Hydrophobic Tail

6-Pentadecanylsalicylic acid, referred to as anacardic acid (C15:0), was found to inhibit the linoleic acid peroxidation competitively catalyzed by soybean lipoxygenase-1 (EC 1.13.11.12, Type 1) with an IC50 of 14.3 μM (4.88 μg/mL). This inhibition is a reversible reaction without pro-oxidant effects. The inhibition kinetics analyzed by Dixon plots indicates that anacardic acid (C15:0) is a competitive inhibitor and the inhibition constant, KI, was established as 6.4 μM. The hydrophilic head (salicylic acid) portion first chelates the iron in the active site and then the hydrophobic tail portion begins reversibly interacting with the C-terminal domain where the iron is located. The inhibition of anacardic acid (C15:0) can be explained by a combination of iron ion-chelation and hydrophobic interaction abilities because of its specific structural feature.

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