The hydrolysis of 6-phosphogluconolactone in the second step of pentose phosphate pathway occurs via a two-water mechanism.

Hydrolysis reaction marks the basis of life yet the mechanism of this crucial biochemical reaction is not completely understood. We recently reported the mechanisms of hydrolysis of nucleoside triphosphate and phosphate monoester. These two reactions hydrolyze P-O-P and P-O-C linkages, respectively. Here, we present the mechanism of hydrolysis of δ-6-phosphogluconolactone, which is an important precursor in the second step of the pentose phosphate pathway. Its hydrolysis requires the cleavage of C-O-C linkage and its mechanism is hitherto unknown. We report three mechanisms of hydrolysis of δ-6-phosphogluconolactone based on density functional computations. In the energetically most favorable mechanism, two water molecules participate in the hydrolysis reaction and the mechanism is sequential, i.e., activation of the attacking water molecule (OH bond breaking) precedes that of the cleavage of the CO bond of the C-O-C linkage. The rate-limiting energy barrier of this mechanism is comparable to the reported experimental free energy barrier. This mechanism has similarities with the mechanism of triphosphate hydrolysis and that of hydrolytic cleavage of DNA in EcoRV enzyme. This two-water sequential hydrolysis mechanism could be the unified mechanism required for the hydrolysis of other hydrolysable species in living cells.

Effect of protonation on the mechanism of phosphate monoester hydrolysis and comparison with the hydrolysis of nucleoside triphosphate in biomolecular motors.

Hydrolysis of phosphate groups is a crucial reaction in living cells. It involves the breaking of two strong bonds, i.e. the OaH bond of the attacking water molecule, and the POl bond of the substrate (Oa and Ol stand for attacking and leaving oxygen atoms). Mechanism of the hydrolysis reaction can proceed either by a concurrent or a sequential mechanism. In the concurrent mechanism, the breaking of OaH and POl bonds occurs simultaneously, whereas in the sequential mechanism, the OaH and POl bonds break at different stages of the reaction. To understand how protonation affects the mechanism of hydrolysis of phosphate monoester, we have studied the mechanism of hydrolysis of protonated and deprotonated phosphate monoester at M06-2X/6-311+G**//M06-2X/6-31+G*+ZPE level of theory (where ZPE stands for zero point energy). Our calculations show that in both protonated and deprotonated cases, the breaking of the water OaH bond occurs before the breaking of the POl bond. Because the two events are not separated by a stable intermediate, the mechanism can be categorized as semi-concurrent. The overall energy barrier is 41kcalmol-1 in the unprotonated case. Most (5/6th) of this is due to the initial breaking of the water OaH bond. This component is lowered from 34 to 25kcalmol-1 by adding one proton to the phosphate. The rest of the overall energy barrier comes from the subsequent breaking of the POl bond and is not sensitive to protonation. This is consistent with previous findings about the effect of triphosphate protonation on the hydrolysis, where the equivalent protonation (on the γ-phosphate) was seen to lower the barrier of breaking the water OaH bond and to have little effect on the POl bond breaking. Hydrolysis pathways of phosphate monoester with initial breaking of the POl bond could not be found here. This is because the leaving group in phosphate monoester cannot be protonated, unlike in triphosphate hydrolysis, where protonation of the ß- and γ-phosphates had been shown to promote a mechanism where the POl bond breaks before the OaH bond does. We also point out that the charge shift due to POl bond breaking during sequential ATP hydrolysis in bio-molecular motors onsets the week unbinding of hydrolysis product that finally leads to the product release during power stroke.

Design, synthesis, in vitro Evaluation and docking studies on dihydropyrimidine-based urease inhibitors.

In our previous report, we have identified 3,4-dihydropyrimidine scaffold as promising class of urease inhibitor in a structure based virtual screen (SBVS) experiment. In present study, we attempted to optimize the scaffold by varying C-5 substituent. The elongation of the C-5 chain was achieved by the reaction of C-5 ester with hydrazine leading to C-5 carbohydrazides which were further used as building blocks for the synthesis of fifteen new compounds having diverse moieties. A significantly higher in vitro urease inhibitory activity with IC50 values in submicromolar range was observed for semithiocarbazide derivatives (4a-c, 0.58-0.79µM) and isatin Schiff base derivative 5a (0.23µM). Docking analysis suggests that the synthesized compounds were anchored well in the catalytic site and extending to the entrance of binding pocket and thus restrict the mobility of the flap by interacting with its key amino acid residues. The overall results of urease inhibition have shown that these compounds can be further optimized and developed as lead urease inhibitors.

Molecular Docking Based Screening of Plant Flavonoids as Dengue NS1 Inhibitors.

Dengue infection has turned into a serious health concern globally due to its high morbidity rate and a high possibility of increase in its mortality rate on the account of unavailability of any proper treatment for severe dengue infection. The situation demands an urgent development of efficient and practicable treatment to deal with Dengue virus (DENV). Flavonoids, a class of phytochemicals present in medicinal plants, possess anti-viral activity and can be strong drug candidates against viruses. NS1 glycoprotein of Dengue virus is involved in its RNA replication and can be a strong target for screening of drugs against this virus. Current study focuses on the identification of flavonoids which can block Asn-130 glycosylation site of Dengue virus NS1 to inhibit viral replication as glycosylation of NS1 is required for its biological functioning. Molecular docking approach was used in this study and the results revealed that flavonoids have strong potential interactions with active site of NS1. Six flavonoids (Deoxycalyxin A; 3,5,7,3',4'-pentahydroxyflavonol-3-O-beta-D-galactopyranoside; (3R)-3',8-Dihydroxyvestitol; Sanggenon O; Epigallocatechin gallate; Chamaejasmin) blocked the Asn-130 glycosylation site of NS1 and could be able to inhibit the viral replication. It can be concluded from this study that these flavonoids could serve as antiviral drugs for dengue infections. Further in-vitro analyses are required to confirm their efficacy and to evaluate their drug potency.

Computer Aided Screening of Phytochemicals from Garcinia against the Dengue NS2B/NS3 Protease.

Dengue virus NS2/NS3 protease because of its ability to cleave viral proteins is considered as an attractive target to screen antiviral agents. Medicinal plants contain a variety of phytochemicals that can be used as drug against different diseases and infections. Therefore, this study was designed to uncover possible phytochemical of different classes (Aromatic, Carbohydrates, Lignin, Saponins, Steroids, Tannins, Terpenoids, Xanthones) that could be used as inhibitors against the NS2B/NS3 protease of DENV. With the help of molecular docking, Garcinia phytochemicals found to be bound deeply inside the active site of DENV NS2B/NS3 protease among all tested phytochemicals and had interactions with catalytic triad (His51, Asp75, Ser135). Thus, it can be concluded from the study that these Gracinia phytochemicals could serve as important inhibitors to inhibit the viral replication inside the host cell. Further in-vitro investigations require confirming their efficacy.

MAPS Database: Medicinal plant Activities, Phytochemical and Structural Database.

UNLABELLED: Drug development from natural sources is an important and fast developing area. Natural sources (plants) have been used to cure a range of diseases for Thousands of years. Different online medicinal plant databases provide information about classifications, activities, phytochemicals and structure of phytochemicals in different formats. These databases do not cover all aspects of medicinal plants. MAPS (Medicinal plant Activities, Phytochemicals & structural database) has been constructed with uniqueness that it combines all information in one web resource and additionally provides test targets on which particular plant found to be effective with reference to the original paper as well. MAPS database is user friendly information resource, including the data of > 500 medicinal plants. This database includes phytochemical constituents, their structure in mol format, different activities possessed by the medicinal plant with the targets reported in literature. AVAILABILITY: http://www.mapsdatabase.com.