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Abstract: Epigallocatechin gallate (EGCG) is known to have numerous pharmacological properties. In the present study, we have shown that EGCG inhibits enoyl–acyl carrier protein reductase of Plasmodium falciparum (PfENR) by following a two-step, slow, tight-binding inhibition mechanism. The association/isomerization rate constant (k 5) of the reversible and loose PfENR–EGCG binary complex to a tight [PfENR–EGCG]∗ or EI∗ complex was calculated to be 4.0×10−2 s−1. The low dissociation rate constant (k 6) of the [PfENR–EGCG]∗ complex confirms the tight-binding nature of EGCG. EGCG inhibited PfENR with the overall inhibition constant (K i ∗) of 7.0±0.8nM. Further, we also studied the effect of triclosan on the inhibitory activity of EGCG. Triclosan lowered the k 6 of the EI∗ complex by 100 times, lowering the overall K i ∗ of EGCG to 97.5±12.5 pM. The results support EGCG as a promising candidate for the development of tea catechin based antimalarial drugs. [Copyright &y& Elsevier]

Abstract: Among the various inhibitors known for enoyl-acyl carrier protein (ACP) reductases, triclosan and green tea catechins are two promising candidates. In the present study, we show, for the first time that epigallocatechin gallate (EGCG), a major component of green tea catechins, inhibits InhA, the enoyl-ACP reductase of Mycobacterium tuberculosis with an IC50 of 17.4μM. EGCG interferes with the binding of NADH to InhA. We also demonstrate that EGCG increased the inhibitory activity of triclosan towards InhA and vice versa. Direct binding assay using [3H]EGCG and fluorescence titration assay support the spectrophotometric/kinetic inhibition data. The biochemical data has been explained by docking simulation studies. [Copyright &y& Elsevier]

Abstract: Acyl carrier protein (ACP) plays a central role in fatty acid biosynthesis by transferring the acyl groups from one enzyme to another for the completion of the fatty acid synthesis cycle. Holo-ACP is the obligatory substrate for the synthesis of acyl-ACPs which act as the carrier and donor for various metabolic reactions. Despite its interactions with numerous proteins in the cell, its mode of interaction is poorly understood. Here, we report the over-expression of PfACP in minimal medium solely in its holo form and in high yield. Expression in minimal media provides a means to isotopically label PfACP for high resolution multi-nuclear and multi-dimensional NMR studies. Indeed, the proton–nitrogen correlated NMR spectrum exhibits very high chemical shift dispersion and resolution. We also show that holo-PfACP thus expressed is amenable to acylation reactions using Escherichia coli acyl-ACP synthetase as well as by standard chemical methods. [Copyright &y& Elsevier]

Abstract: A gamma-kafirin gene of Sorghum bicolor L var. M35-1 was isolated using a PCR based approach by designing specific primers. The primers gave highly reproducible amplification. The amplicons were then cloned and sequenced. Nucleotide sequences were subjected to the homology search and a comparative analysis was done with other prolamins. Amplified γ-kafirin gene (accession no. AY566298–99 AY566298 AY566299 ) showed 99% homology with a γ-kafirin gene of Sorghum vulgare. Compared to Sorghum vulgare, only 3 extra bases were present in Sorghum bicolor at position 40 nucleotide bases downstream of transcription initiation site. These sequences were related with prolamins of other genera, i.e. coix and maize with 84% sequence homology. The deduced protein sequence of γ-kafirin gene of S. bicolor (accession no. AAS73290) gave significant similarity of 99%, 79% and 77% with gamma-kafirin protein of S. vulgare, γ-zein and γ-coixin proteins, respectively. All cysteine residues of γ-kafirin were highly conserved. Probable secondary structure of gamma-kafirin protein was predicted using the online PSIPRED server. Study of temporal expression of the γ-kafirin gene is needed for analysing the expression pattern of introgressed gene(s) driven by its promoter. Temporal expression of gamma-kafirin gene in endosperm studied by semi quantitative RT–PCR with specifically designed primers showed that γ-kafirin expression started at 7DAP (days after pollination). There was statistically a non-significant increase in expression up to 14DAP, thereafter expression increased significantly (at the 5% level) and reached a maximum at 21DAP. Expression of γ-kafirin started decreasing after 21DAP and a very low expression was detected at 28DAP. [Copyright &y& Elsevier]