ABSTRACT
In the search for new antiparasitic natural compounds from the medicinal plants from Cameroon, the new 22-hydroxyclerosterol, established as such on the basis of detailed chemical and spectroscopic analysis, was isolated from the hexane extract of the stem bark of Allexis cauliflora together with the known clerosterol. 22-Hydroxyclerosterol inhibited the growth of Trypanosoma brucei brucei cells with an ED(50) value of 1.56 µM. The compound was also established as an uncompetitive inhibitor of the glycolytic enzyme PGI of T. brucei (Ki'= 3 ± 1 µM), an uncompetitive inhibitor of mammalian rabbit muscles' enzyme PyK (Ki'= 26 ± 3 µM) and a mixed inhibitor of PyK of Leishmania mexicana (Ki'= 65 ± 10 µM; Ki= 24 ± 5 µM).
ABSTRACT
In continuation of our study on medicinal plants of Cameroon, stem barks of Polyalthia suaveolens were phytochemically studied. This investigation yielded a new indolosesquiterpene alkaloid, named polysin (1) and four hitherto known alkaloids (2-5). Polysin (1) appeared as a competitive reversible inhibitor (K(i)=10 microM) of phosphofructo kinase (PFK) of Trypanosoma brucei with respect to fructose-6-phosphate (K(i)/K(M)=0.05) and could be used in the design of new trypanocidal drugs. The other isolated compounds (2-5) also exhibited interesting inhibitory effects on selected glycolytic enzymes (PFK, glyceraldehyde-3-phosphate dehydrogenase and aldolase).
Subject(s)
Alkaloids/pharmacology , Polyalthia/chemistry , Trypanocidal Agents/pharmacology , Trypanosoma brucei brucei/enzymology , Alkaloids/chemistry , Alkaloids/isolation & purification , Cameroon , Enzyme Inhibitors/pharmacology , Glycolysis/drug effects , Phosphofructokinases/antagonists & inhibitors , Phytotherapy , Plants, Medicinal/chemistry , Sesquiterpenes , Trypanocidal Agents/chemistry , Trypanocidal Agents/isolation & purification , Trypanosoma brucei brucei/drug effects , Trypanosoma brucei brucei/metabolismABSTRACT
In an attempt to establish the components responsible for the use of Enantia chlorantha against cutaneous leishmaniasis in local traditional medicine, a well-known palmatine has been isolated in substantial amounts from a methanolic bark extract of this plant species. Palmatine therein obtained exhibited a significant inhibitory activity on growth of both Trypanosoma cruzi (IC(50) 0.068 microM) and Leishmania infantum (IC(50) 0.79 microM).
Subject(s)
Annonaceae/chemistry , Antiprotozoal Agents/pharmacology , Berberine Alkaloids/pharmacology , Leishmania infantum/drug effects , Plant Extracts/pharmacology , Trypanosoma cruzi/drug effects , Animals , Antiprotozoal Agents/chemistry , Antiprotozoal Agents/isolation & purification , Berberine Alkaloids/chemistry , Berberine Alkaloids/isolation & purification , Plant Bark/chemistry , Plant Extracts/chemistry , Plant Extracts/isolation & purification , Trypanocidal Agents/chemistry , Trypanocidal Agents/isolation & purification , Trypanocidal Agents/pharmacologyABSTRACT
Polycarpol, sitosterol and sitosterol-3-O-beta-D-glucoside isolated for the first time from Piptostigma preussi (Annonaceae) occur regularly in some Annonaceae such as Piptostigma genus. Polycarpol exhibits interesting antitrypanosomal activity with an ED(50) value of 5.11 microM on Trypanosoma brucei cells. Moreover, it inhibits T. brucei glycolytic enzymes GAPDH and PFK with IC(50) values of 650 and 180 microM respectively.
Subject(s)
Annonaceae/chemistry , Plant Extracts/pharmacology , Polycyclic Aromatic Hydrocarbons/pharmacology , Trypanocidal Agents/pharmacology , Trypanosoma brucei brucei/drug effects , Enzyme Inhibitors/isolation & purification , Enzyme Inhibitors/pharmacology , Glucosides/isolation & purification , Plant Bark , Plant Extracts/chemistry , Plant Stems , Polycyclic Aromatic Hydrocarbons/isolation & purification , Sitosterols/isolation & purification , Trypanocidal Agents/isolation & purification , Trypanosoma brucei brucei/enzymologyABSTRACT
A new series of "bi-substrate" analogues have been synthesized as potential inhibitors of the glyceraldehyde-3-phosphate dehydrogenase and one lead compound has been identified that inhibits the enzyme from Trypanosoma cruzi with good affinity and very high (50-fold) specificity.
Subject(s)
Enzyme Inhibitors/pharmacology , Glyceraldehyde-3-Phosphate Dehydrogenases/antagonists & inhibitors , Trypanosoma cruzi/enzymology , AnimalsABSTRACT
We report here the first crystal structure of a stable isosteric analogue of 1,3-bisphospho-d-glyceric acid (1,3-BPGA) bound to the catalytic domain of Trypanosoma cruzi glycosomal glyceraldehyde-3-phosphate dehydrogenase (gGAPDH) in which the two phosphoryl moieties interact with Arg249. This complex possibly illustrates a step of the catalytic process by which Arg249 may induce compression of the product formed, allowing its expulsion from the active site. Structural modifications were introduced into this isosteric analogue and the respective inhibitory effects of the resulting diphosphorylated compounds on T. cruzi and Trypanosoma brucei gGAPDHs were investigated by enzymatic inhibition studies, fluorescence spectroscopy, site-directed mutagenesis, and molecular modelling. Despite the high homology between the two trypanomastid gGAPDHs (> 95%), we have identified specific interactions that could be used to design selective irreversible inhibitors against T. cruzi gGAPDH.
Subject(s)
Diphosphoglyceric Acids/metabolism , Drug Design , Glyceraldehyde 3-Phosphate Dehydrogenase (NADP+)/chemistry , Glyceraldehyde 3-Phosphate Dehydrogenase (NADP+)/metabolism , Trypanosoma cruzi/enzymology , Animals , Binding Sites , Crystallography, X-Ray , Diphosphoglyceric Acids/chemistry , Diphosphoglyceric Acids/pharmacology , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/metabolism , Enzyme Inhibitors/pharmacology , Glyceraldehyde 3-Phosphate Dehydrogenase (NADP+)/antagonists & inhibitors , Kinetics , Models, Molecular , Molecular Conformation , Protein Binding , Protein Structure, Tertiary , Species Specificity , Static Electricity , Structure-Activity Relationship , Trypanosoma brucei brucei/enzymologyABSTRACT
This work deals with the phosphofructokinase enzyme (PFK) of the parasite Trypanosoma brucei. Inhibitors which are analogues of fructose-6-phosphate (F6P) derived from 2,5-anhydromannitol and therefore blocked in a closed conformation, both nonphosphorylated and phosphorylated, were designed. They provided information on this class of ATP-dependent PFK (structurally more similar to PPi-dependent PFKs revealing (i) an ordered mechanism, ATP binding first, inducing an essential conformational change to increase the affinity for F6P, and (ii) a rather hydrophobic environment at the ATP binding site. Nonphosphorylated mannitol derivatives bind at both the ATP and F6P binding sites, whereas the phosphorylated derivatives only bind at the ATP binding site. The inhibitors bearing an aromatic ring substituted at the meta position indicate a polar interaction with lysine 227, which is specific to T. brucei PFK and is replaced by a glycine in human PFK. This lysine can be irreversibly bound, leading to inhibition when an electrophilic carbon atom is beta to the meta position on the ring. This lysine was identified by site-directed mutagenesis. This first example of a specific irreversible inactivation of T. brucei PFK offers an opportunity to develop biologically active compounds against the sleeping sickness, the causative agent of which is the trypanosome.
Subject(s)
Phosphofructokinase-1/antagonists & inhibitors , Phosphofructokinase-1/metabolism , Trypanosoma brucei brucei/enzymology , Adenosine Triphosphate/chemistry , Adenosine Triphosphate/metabolism , Animals , Binding Sites , Binding, Competitive , Circular Dichroism , Enzyme Activation/drug effects , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/pharmacology , Fructosephosphates/chemistry , Fructosephosphates/metabolism , Kinetics , Magnesium/chemistry , Phosphofructokinase-1/chemistry , Recombinant Proteins/antagonists & inhibitors , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Spectrometry, Fluorescence/methods , Substrate SpecificityABSTRACT
BACKGROUND: In humans, there are four alkaline phosphatases, and each form exhibits a characteristic pattern of tissue distribution. The availability of an easy method to reveal their activity has resulted in large amount of data reporting correlations between variations in activity and illnesses. For example, alkaline phosphatase from neutrophils of mothers pregnant with a trisomy 21 fetus (Down's syndrome) displays significant differences both in its biochemical and immunological properties, and in its affinity for some specific inhibitors. RESULTS: To analyse these differences, the biochemical characteristics of two isozymes (non specific and placental alkaline phosphatases) were expressed in baculovirus infected cells. Comparative analysis of the two proteins allowed us to estimate the kinetic constants of denaturation and sensitivity to two inhibitors (L-p-bromotetramisole and thiophosphate), allowing better discrimination between the two enzymes. These parameters were then used to estimate the ratio of the two isoenzymes in neutrophils of pregnant mothers with or without a trisomy 21 fetus. It appeared that the placental isozyme represented 13% of the total activity of neutrophils of non pregnant women. This proportion did not significantly increase with normal pregnancy. By contrast, in pregnancies with trisomy 21 fetus, the proportion reached 60-80% of activity. CONCLUSION: Over-expression of the placental isozyme compared with the tissue-nonspecific form in neutrophils of mother with a trisomy 21 fetus may explain why the characteristics of the alkaline phosphatase in these cells is different from normal. Application of this knowledge could improve the potential of using alkaline phosphatase measurements to screen for Down's syndrome.
