Anti-amyloid potential of some phytochemicals against Aß-peptide and α-synuclein, tau, prion, and Huntingtin protein.

Some molecules self-assemble to create complex structures through molecular self-assembly. Hydrogel preparation, tissue repair, and therapeutic drug delivery are a few applications of molecular self-assembly. However, the self-assembly of amino acids, peptides, and proteins forms amyloid fibrils, resulting in various disorders, most notably neurodegenerative ailments. Examples include the self-assembly of phenylalanine, which causes phenylketonuria; Aß, which causes Alzheimer's disease; the tau protein, which causes both Alzheimer's and Parkinson's diseases; and α-synuclein, which causes Parkinson's illness. This review provides information related to phytochemicals of great significance that can prevent the formation of, or destabilize, amino acid, peptide, and protein self-assemblies.

The self-assembly of L-histidine might be the cause of histidinemia.

L-Histidine is an essential amino acid with unique biochemical and physiological properties. Histidinemia is a disease condition caused by the elevated level of L-histidine in our blood. Mutations in the histidase, an enzyme for the breakdown of histidine, is the cause of the rise in histidine concentration. To our knowledge, no research has been done on why a high concentration of histidine causes histidinemia. In this study, we provide a potential explanation why the elevated levels of histidine in the human body causes histidinemia. In this study we have found that L-histidine self-assembled in water to form nano sheet structures at physiological pH and temperature, using 1D 1H NMR spectroscopy, diffusion ordered spectroscopy (DOSY) and scanning electron microscope (SEM) techniques. The kinetics of self-assembly has been studied using real time NMR spectroscopy. We observed that both the aromatic ring and aliphatic part are equally contributing to the self-assembly of L-histidine. The symptoms of histidinemia, neurological deficits and speech delays, are similar to that of the neurodegenerative diseases caused by the self-assembly of peptides and proteins. We speculate that the self-assembly of L-histidine might be the cause of histidinemia.

Acute promyelocytic leukemia drug - arsenic trioxide in the presence of eugenol shows differential action on leukemia cells (HL-60) and cardiomyocytes (H9c2) - inference from NMR study.

The increased concern of cardiovascular dysfunction by cancer therapeutics has led to more effective treatment strategies. Arsenic trioxide (As2O3) is a potential chemotherapeutic agent for acute promyelocytic leukemia (APL), but the effectiveness is affected by potential cardiotoxicity. Researchers have been trying to find out novel modalities to manage the adverse effects of As2O3. In our study, the antioxidant molecule eugenol showed protective action against the destructive impact of As2O3 on cardiomyocytes (H9c2) without compromising the anti-cancer property As2O3 on leukemia cells (HL-60). We have studied the interaction between arsenic and eugenol in physiological and acidic pH to understand the molecular mechanism of differential action of As2O3 in the presence of eugenol using NMR spectroscopy. The study observed that at physiological pH, arsenic and eugenol interact to form an inactive product, positively affecting H9c2 cardiomyocytes. Still, there is no such interaction in acidic pH evidenced by the useful anti-cancer property of As2O3. The result concludes that the antioxidant molecule eugenol is an efficient protective agent against the adverse effect of As2O3 on cardiomyocytes.

Pressure response of 31P chemical shifts of adenine nucleotides.

High pressure NMR spectroscopy is a powerful method for identifying rare conformational states of proteins from the pressure response of their chemical shifts. Many proteins have bound adenine nucleotides at their active centers, in most cases in a complex with Mg2+-ions. The 31P NMR signals of phosphate groups of the nucleotides can be used as probes for conformational transitions in the proteins themselves. For distinguishing protein specific pressure effects from trivial pressure responses not due to the protein interaction, data of the pressure response of the free nucleotides must be available. Therefore, the pressure response of 31P chemical shifts of the adenine nucleotides AMP, ADP, and ATP and their Mg2+-complexes has been determined at pH values several units distant from the respective pK-values. It is clearly non-linear for most of the resonances. A negative first order pressure coefficient B1 was determined for all 31P resonances except Mg2+·AMP indicating an upfield shift of the resonances with pressure. The smallest and largest negative values are obtained for the α-phosphate group of ADP and ß-phosphate group of Mg2+·ATP with -0.32 and -4.59ppm/GPa, respectively. With exception of the α-phosphate group of Mg2+·AMP the second order pressure coefficients are positive leading to a saturation like behaviour. The pressure response of the adenine nucleotides is similar but not identical to that observed earlier for guanine nucleotides. The obtained data show that the chemical shift pressure response of the different phosphate groups is rather different dependent on the position of phosphate group in the nucleotide and the nucleotide used.

Structural transitions in full-length human prion protein detected by xenon as probe and spin labeling of the N-terminal domain.

Fatal neurodegenerative disorders termed transmissible spongiform encephalopathies (TSEs) are associated with the accumulation of fibrils of misfolded prion protein PrP. The noble gas xenon accommodates into four transiently enlarged hydrophobic cavities located in the well-folded core of human PrP(23-230) as detected by [(1)H, (15)N]-HSQC spectroscopy. In thermal equilibrium a fifth xenon binding site is formed transiently by amino acids A120 to L125 of the presumably disordered N-terminal domain and by amino acids K185 to T193 of the well-folded domain. Xenon bound PrP was modelled by restraint molecular dynamics. The individual microscopic and macroscopic dissociation constants could be derived by fitting the data to a model including a dynamic opening and closing of the cavities. As observed earlier by high pressure NMR spectroscopy xenon binding influences also other amino acids all over the N-terminal domain including residues of the AGAAAAGA motif indicating a structural coupling between the N-terminal domain and the core domain. This is in agreement with spin labelling experiments at positions 93 or 107 that show a transient interaction between the N-terminus and the start of helix 2 and the end of helix 3 of the core domain similar to that observed earlier by Zn(2+)-binding to the octarepeat motif.

Sequential backbone resonance assignments of the E. coli dihydrofolate reductase Gly67Val mutant: folate complex.

Occasionally, a mutation in an exposed loop region causes a significant change in protein function and/or stability. A single mutation Gly67Val of E. coli dihydrofolate reductase (DHFR) in the exposed CD loop is such an example. We have carried out the chemical shift assignments for H(N), N(H), C(α) and C(ß) atoms of the Gly67Val mutant of E. coli DHFR complexed with folate at pH 7.0, 35 °C, and then evaluated the H(N), N(H), C(α) and C(ß) chemical shift changes caused by the mutation. The result indicates that, while the overall secondary structure remains the same, the single mutation Gly67Val causes site-specific conformational changes of the polypeptide backbone restricted around the adenosine-binding subdomain (residues 38-88) and not in the distant catalytic domain.

Functional conformer of c-Myb DNA-binding domain revealed by variable temperature studies.

The conformational fluctuation in the minimum DNA-binding domain of c-Myb, repeats 2 and 3 (R2R3), was studied under closely physiological conditions. A global unfolding transition, involving both the main chain and the side chains, was found to take place at the approximate temperature range 30-70 °C, with a transition temperature of approximately 50 °C. In addition, the observation of simultaneous shift change and broadening of NMR signals in both (1)H one-dimensional and (15)N/(1)H two-dimensional NMR spectra indicated the occurrence of locally fluctuating state at physiological temperature. In the wild-type protein containing a cavity in R2, the local fluctuation of R2 is more prominent than that of R3, whereas it is suppressed in the cavity-filled mutant, V103L. This indicates that the cavity in R2 contributes significantly to the conformational instability and the transition into the locally fluctuating state. For the wild-type R2R3 protein, the more dynamic conformer is estimated to be present to some extent at 37 °C and is likely beneficial for its biological function: DNA-binding. This result is in agreement with the concept of an excited-state conformer that exists in equilibrium with the dominant ground-state conformer and acts as the functional conformer of the protein. From the findings of the present study, it appears that the tandem repeats of two small domains with no disulfide bonds and with a destabilizing cavity function as the evolutionary strategy of the wide-type c-Myb DNA-binding domain to produce an appropriate fraction of the locally fluctuating state at 37 °C, which is more amenable to DNA-binding. Database: Chemical shifts and peak lists have been deposited in the Biological Magnetic Resonance Bank under entries 11584 and 11585.

Interactions of some commonly used drugs with human α-thrombin.

Adverse side effects of drugs are often caused by the interaction of drug molecules to targets other than the intended ones. In this study, we investigated the off-target interactions of some commercially available drugs with human α-thrombin. The drugs used in the study were selected from Super Drug Database based on the structural similarity to a known thrombin inhibitor argatroban. Interactions of these drugs with thrombin were initially checked by in silico docking studies and then confirmed by thrombin inhibition assay using a fluorescence microplate-based method. Results show that the three commonly used drugs piperacillin (anti-bacterial), azlocillin (anti-bacterial), and metolazone (anti-hypertensive and diuretic) have thrombin inhibitory activity almost similar to that of argatroban. The Ki values of piperacillin, azlocillin, and metolazone with thrombin are .55, .95, and .62 nM, respectively. The IC50 values of piperacillin, azlocillin, and metolazone with thrombin are 1.7, 2.9, and 1.92 nM, respectively. This thrombin inhibitory activity might be a reason for the observed side effects of these drugs related to blood coagulation and other thrombin activities. Furthermore, these compounds (drugs) may be used as anti-coagulants as such or with structural modifications.

In silico characterization of a novel ß-1,3-glucanase gene from Bacillus amyloliquefaciens--a bacterial endophyte of Hevea brasiliensis antagonistic to Phytophthora meadii.

We report the molecular characterization of ß-1,3-glucanase-producing Bacillus amyloliquefaciens-an endophyte of Hevea brasiliensis antagonistic to Phytophthora meadii. After cloning and sequencing, the ß-1,3-glucanase gene was found to be 747 bp in length. A homology model of the ß-1,3-glucanase protein was built from the amino acid sequence obtained upon translation of the gene. The target ß-1,3-glucanase protein and the template protein, endo ß-1,3-1,4-glucanase protein (PDB ID: 3o5s), were found to share 94% sequence identity and to have similar secondary and tertiary structures. In the modeled structure, three residues in the active site region of the template-Asn52, Ile157 and Val158-were substituted with Asp, Leu and Ala, respectively. Computer-aided docking studies of the substrate disaccharide (ß-1, 3-glucan) with the target as well as with the template proteins showed that the two protein-substrate complexes were stabilized by three hydrogen bonds and by many van der Waals interactions. Although the binding energies and the number of hydrogen bonds were the same in both complexes, the orientations of the substrate in the active sites of the two proteins were different. These variations might be due to the change in the three amino acids in the active site region of the two proteins. The difference in substrate orientation in the active site could also affect the catalytic potential of the ß-1,3 glucanase enzyme.