Theoretical Study on Zigzag Boron Nitride Nanowires.

In this work, two kinds of BN-nanowires (BNnws): a-BNnw and d-BNnw, respectively composed of azo (N-N) and diboron (B-B) bonds, are proposed with the aid of the first-principles simulations. Their structural stabilities are carefully verified from the energetics, lattice dynamics, and thermodynamic perspectives. Similar to the other common boron nitride polymorph, the a-BNnw and d-BNnw are semiconductors with relatively wide band gaps of 3.256 and 4.631 eV at the HSE06 level, respectively. The corresponding projected DOS patterns point out that their band edges are composed of different atomic species, which can help with the separation of their excitons. The band gaps can be manipulated monotonically by axial strains within the elastic ranges. The major charge carriers are electron holes. Significantly, a-BNnw possesses very high carrier mobilities around 0.44×104  cm2 V-1 s-1 .

Enhancing the potency of lithospermate B for inhibiting Na+/K+-ATPase activity by forming transition metal ion complexes.

AIM: To determine whether replacing Mg(2+) in magnesium lithospermate B (Mg-LSB) isolated from danshen (Salvia miltiorrhiza) with other metal ions could affect its potency in inhibition of Na(+)/K(+)-ATPase activity. METHODS: Eight metal ions (Na(+), K(+), Mg(2+), Cr(3+), Mn(2+), Co(2+), Ni(2+), and Zn(2+)) were used to form complexes with LSB. The activity of Na(+)/K(+)-ATPase was determined by measuring the amount of inorganic phosphate (Pi) liberated from ATP. Human adrenergic neuroblastoma cell line SH-SY5Y was used to assess the intracellular Ca(2+) level fluctuation and cell viability. The metal binding site on LSB and the binding mode of the metal-LSB complexes were detected by NMR and visible spectroscopy, respectively. RESULTS: The potencies of LSB complexed with Cr(3+), Mn(2+), Co(2+), or Ni(2+) increased by approximately 5 times compared to the naturally occurring LSB and Mg-LSB. The IC50 values of Cr-LSB, Mn-LSB, Co-LSB, Ni-LSB, LSB, and Mg-LSB in inhibition of Na(+)/K(+)-ATPase activity were 23, 17, 26, 25, 101, and 128 µmol/L, respectively. After treatment of SH-SY5Y cells with the transition metal-LSB complexes (25 µmol/L), the intracellular Ca(2+) level was substantially elevated, and the cells were viable for one day. The transition metals, as exemplified by Co(2+), appeared to be coordinated by two carboxylate groups and one carbonyl group of LSB. Titration of LSB against Co(2+) demonstrated that the Co-LSB complex was formed with a Co(2+):LSB molar ratio of 1:2 or 1:1, when [Co(2+)] was less than half of the [LSB] or higher than the [LSB], respectively. CONCLUSION: LSB complexed with Cr(3+), Mn(2+), Co(2+), or Ni(2+) are stable, non-toxic and more potent in inhibition of Na(+)/K(+)-ATPase. The transition metal-LSB complexes have the potential to be superior substitutes for cardiac glycosides in the treatment of congestive heart failure.

Site specificity of OH α-H abstraction reaction for a ß-hairpin peptide: an ab initio study.

A ß-hairpin peptide (PDB ID 1UAO) was modeled to explore the backbone oxidation of a protein by an OH radical to abstract one α-H atom with ab initio calculation at the B3LYB/6-31G(d) without any constraint. Three glycine residues located at three different sites in 1UAO were used to examine the possible site specificity of this backbone oxidation. The pre- and post-reactive complexes along with their associated transition states were located and verified by the intrinsic reaction coordinate method. The reaction profile of these α-H abstraction reactions was constructed. The effects of the aqueous solution were estimated by the conductor-like polarizable continuum model (CPCM) model. Rate constants were calculated with transition state theory. The reaction rate of the OH α-H abstraction varies among these three different sites. The differences among these three sites were rationalized in terms of the molecular and electronic structures of the reactive complexes along the reaction pathway. The explicit solvation effect was estimated through the similar abstraction of a zwitterionic glycine with the combination of microsolvation and a CPCM model. Our results indicate that the α-H abstraction at certain sites requires explicit salvation to obtain accurate results. A replica exchange molecular dynamics simulation was performed to demonstrate the structural change due to this type of abstraction.

Antcin A, a steroid-like compound from Antrodia camphorata, exerts anti-inflammatory effect via mimicking glucocorticoids.

AIM: To determine the active ingredient of Niuchangchih (Antrodia camphorata) responsible for its anti-inflammatory effects and the relevant molecular mechanisms. METHODS: Five major antcins (A, B, C, H, and K) were isolated from fruiting bodies of Niuchangchih. Structural similarity between the antcins and 2 glucocorticoids (cortisone and dexamethasone) was compared. After incubation with each compound, the cytosolic glucocorticoid receptor (GR) was examined for its migration into the nucleus. Mo lecular docking was performed to model the tertiary structure of GR associated with antcins. RESULTS: Incubation with cortisone, dexamethasone or antcin A (but not antcins B, C, H, and K) led to the migration of glucocorticoid receptor into the nucleus. The minimal concentration of antcin A, cortisone and dexamethasone to induce nuclear migration of glucocorticoid receptor was 10, 1, and 0.1 mol/L, respectively. The results are in agreement with the simulated binding affinity scores of these three ligands docking to the glucocorticoid receptor. Molecular modeling indicates that C-7 of antcin A or glucocorticoids is exposed to a hydrophobic region in the binding cavity of the glucocorticoid receptor, and the attachment of a hydrophilic group to C-7 of the other four antcins presumably results in their being expelled when docking to the cavity. CONCLUSION: The anti-inflammatory effect of Niuchangchih is, at least, partly attributed to antcin A that mimics glucocorticoids and triggers translocation of glucocorticoid receptor into nucleus to initiate the suppressing inflammation.

Docking study of the precursor peptide of mastoparan onto its putative processing enzyme, dipeptidyl peptidase IV: a revisit to molecular ticketing.

Stepwise-cleavage process of promastoparans to reach maturity was investigated theoretically by combining ab initio folding and unbounded docking. The comparison between the structures of the promastoparans both before and after docking were examined along with the hydrogen bonding interaction pattern between the dipetidyl peptidase IV (DPPIV) and promastoparans to reveal how the endpoint of this stepwise cleavage is recognized among these promastoparans with highly resemble amino acid sequences. The current approach of folding and docking study provides structural insight on the stepwise cleavage process.

Magnesium lithospermate B extracted from Salvia miltiorrhiza elevates intracellular Ca(2+) level in SH-SY5Y cells.

AIM: To examine if magnesium lithospermate B (MLB), a potent inhibitor of Na(+)/K(+)-ATPase, leads to the elevation of intracellular Ca(2+) level as observed in cells treated with cardiac glycosides. METHODS: Viability of SH-SY5Y neuroblastoma cells treated with various concentrations of ouabain or MLB was measured. Intracellular Ca(2+) levels were visualized using Fluo4-AM (fluorescent dye) when cells were treated with ouabain or MLB in the presence or absence of KB-R7943 (Na(+)/Ca(2+) exchanger inhibitor) and 2-APB (IP(3) receptor antagonist). Molecular modeling was conducted for the docking of ouabain or MLB to Na(+)/K(+)-ATPase. Changes of cell body and dendrite morphology were monitored under a microscope. RESULTS: severe toxicity was observed in cells treated with ouabain of concentration higher than 1 micromol/L for 24 h while no apparent toxicity was observed in those treated with MLB. Intracellular Ca(2+) levels were substantially elevated by MLB (1 micromol/L) and ouabain (1 micromol/L) in similar patterns, and significantly reduced in the presence of KB-R7943 (10 micromol/L) or 2-APB (100 micromol/L). Equivalent interaction with the binding cavity of Na(+)/K(+)-ATPase was simulated for ouabain and MLB by forming five hydrogen bonds, respectively. Treatment of ouabain (1 micromol/L), but not MLB (1 mumol/L), induced dendritic shrink of SH-SY5Y cells. CONCLUSION: Comparable to ouabain, MLB leads to the elevation of intracellular Ca(2+) level presumably via the same mechanism by inhibiting Na(+)/K(+)-ATPase. The elevated Ca(2+) levels seem to be supplied by Ca(2+) influx through the reversed mode of the Na(+)/Ca(2+) exchanger and intracellular release from endoplasmic reticulum.

Steroid-like compounds in Chinese medicines promote blood circulation via inhibition of Na+/K+ -ATPase.

AIM: To examine if steroid-like compounds found in many Chinese medicinal products conventionally used for the promotion of blood circulation may act as active components via the same molecular mechanism triggered by cardiac glycosides, such as ouabain. METHODS: The inhibitory potency of ouabain and the identified steroid-like compounds on Na(+)/K(+)-ATPase activity was examined and compared. Molecular modeling was exhibited for the docking of these compounds to Na(+)/K(+)-ATPase. RESULTS: All the examined steroid-like compounds displayed more or less inhibition on Na(+)/K(+)-ATPase, with bufalin (structurally almost equivalent to ouabain) exhibiting significantly higher inhibitory potency than the others. In the pentacyclic triterpenoids examined, ursolic acid and oleanolic acid were moderate inhibitors of Na(+)/K(+)-ATPase, and their inhibitory potency was comparable to that of ginsenoside Rh2. The relatively high inhibitory potency of ursolic acid or oleanolic acid was due to the formation of a hydrogen bond between its carboxyl group and the Ile322 residue in the deep cavity close to two K(+) binding sites of Na(+)/K(+)-ATPase. Moreover, the drastic difference observed in the inhibitory potency of ouabain, bufalin, ginsenoside Rh2, and pentacyclic triterpenoids is ascribed mainly to the number of hydrogen bonds and partially to the strength of hydrophobic interaction between the compounds and residues around the deep cavity of Na(+)/K(+)-ATPase. CONCLUSION: Steroid-like compounds seem to contribute to therapeutic effects of many cardioactive Chinese medicinal products. Chinese herbs, such as Prunella vulgaris L, rich in ursolic acid, oleanolic acid and their glycoside derivatives may be adequate sources for cardiac therapy via effective inhibition on Na(+)/K(+)-ATPase.

Site specificity of the (alpha)C--H bond dissociation energy for a naturally occurring beta-hairpin peptide-An ab initio study.

A naturally occurring beta-hairpin peptide (PDB ID 1UAO) was used as a model to study the backbone oxidation of a protein with ab initio calculation at the B3LYB/6-31G(d) without any constraints. The (alpha)C--H bond dissociation energy of three different glycyl radicals located at different sites on the beta-hairpin peptide was calculated to evaluate the site specificity of backbone oxidation. The molecular and electronic structures of these glycyl radicals were analyzed to rationalize this site specificity. The overall molecular structure of the alpha-H abstracted beta-hairpin peptide remained almost unchanged with the exception of the local conformation of the attacked residue. However, the (alpha)C--H bond strength varied dramatically among these different sites.

Energy landscapes associated with the self-aggregation of an alanine-based oligopeptide (AAKA)4.

The alanine-based 16-mer peptide (AAKA)(4) has recently been shown to aggregate into a hydrogel at centimolar concentrations and high ionic strength ( Measey, T. J.; Schweitzer-Stenner, R. J. Am. Chem. Soc. 2006 , 128 , 13324. ). This is a surprising result since the current understanding of peptide self-aggregation would lead one to expect that the positively charged lysine residues inhibit the formation of beta-sheet structures. The present study was aimed at shedding some light on the mechanism governing the initial phase of the self-aggregation process. To this end we measured CD and FTIR spectra of the (AAKA)(n) at millimolar concentration and low ionic strength and found that the peptide forms soluble aggregates rather than a hydrogel under these conditions. To analyze the initial phase of this aggregation process, we carried out simulations on the dimerization and trimerization of (AAKA)(4) using replica exchange molecular dynamics (MD) methods based on an implicit solvent model. The results indicate that the peptide aggregates into stable antiparallel beta-sheet structures under conditions comparable to experiments. Furthermore, the existence of trimers is a very sensitive function of the peptide concentration and temperature; no significant amount of trimers is formed at low concentration and the trimer population drops sharply as temperature increases above about 320 K. These results on oligomers are consistent with the trend observed in our experiments (reported elsewhere). We have also obtained the free energy landscapes of the trimers as a function of reaction coordinates, which reveal a few basins of minimal free energy. Associated with them, six major types of stable conformers were identified and the structures of transition states between them were also determined. We suggest a possible transition mechanism between these conformers based on the landscape and structural analysis of the aggregates.

Effect of sugar positions in ginsenosides and their inhibitory potency on Na+/K+-ATPase activity.

AIM: To determine whether ginsenosides with various sugar attachments may act as active components responsible for the cardiac therapeutic effects of ginseng and sanqi (the roots of Panax ginseng and Panax notoginseng) via the same molecular mechanism triggered by cardiac glycosides, such as ouabain and digoxin. METHODS: The structural similarity between ginsenosides and ouabain was analyzed. The inhibitory potency of ginsenosides and ouabain on Na+/K+-ATPase activity was examined and compared. Molecular modeling was exhibited for the docking of ginsenosides to Na+/K+-ATPase. RESULTS: Ginsenosides with sugar moieties attached only to the C-3 position of the steroid-like structure, equivalent to the sugar position in cardiac glycosides, and possessed inhibitory potency on Na+/K+-ATPase activity. However, their inhibitory potency was significantly reduced or completely abolished when a monosaccharide was linked to the C-6 or C-20 position of the steroid-like structure; replacement of the monosaccharide with a disaccharide molecule at either of these positions caused the disappearance of the inhibitory potency. Molecular modeling and docking confirmed that the difference in Na+/K+-ATPase inhibitory potency among ginsenosides was due to the steric hindrance of sugar attachment at the C-6 and C-20 positions of the steroid-like structure. CONCLUSION: The cardiac therapeutic effects of ginseng and sanqi should be at least partly attributed to the effective inhibition of Na+/K+-ATPase by their metabolized ginsenosides with sugar moieties attached only to the C-3 position of the steroid-like structure.

Construction and comparison of fluorescence and bioluminescence bacterial biosensors for the detection of bioavailable toluene and related compounds.

Environmental pollution with petroleum products such as benzene, toluene, ethylbenzene, and xylenes (BTEX) has garnered increasing awareness because of its serious consequences for human health and the environment. We have constructed toluene bacterial biosensors comprised of two reporter genes, gfp and luxCDABE, characterized by green fluorescence and luminescence, respectively, and compared their abilities to detect bioavailable toluene and related compounds. The bacterial luminescence biosensor allowed faster and more-sensitive detection of toluene; the fluorescence biosensor strain was much more stable and thus more applicable for long-term exposure. Both luminescence and fluorescence biosensors were field-tested to measure the relative bioavailability of BTEX in contaminated groundwater and soil samples. The estimated BTEX concentrations determined by the luminescence and fluorescence bacterial biosensors were closely comparable to each other. Our results demonstrate that both bacterial luminescence and fluorescence biosensors are useful in determining the presence and the bioavailable fractions of BTEX in the environment.

Theoretical investigation of the photoinitiated folding of HP-36.

A computational model was developed to examine the phototriggered folding of a caged protein, a protein modified with an organic photolabile cross-linker. Molecular dynamics simulations of the modified 36-residue fragment of subdomain B of chicken villin head piece with a photolabile linker were performed, starting from both the caged and the uncaged structures. Construction of a free-energy landscape, based on principal components as well as on radius of gyration versus root-mean-square deviation, and circular dichroism calculations were employed to characterize folding behavior and structures. The folded structures observed in the molecular dynamics trajectories were found to be similar to that of the wild-type protein, in agreement with the published experimental results. The free-energy landscapes of the modified and wild-type proteins have similar topology, suggesting common thermodynamic/kinetic behavior. The existence of small differences in the free-energy surface of the modified protein from that of the native protein, however, indicates subtle differences in the folding behavior.

Gene families encoding isoforms of two major sesame seed storage proteins, 11S globulin and 2S albumin.

Sesame (Sesamum indicum L.) seed has been recognized as a nutritional protein source owing to its richness in methionine. Storage proteins have been implicated in allergenic responses to sesame consumption. Two abundant storage proteins, 11S globulin and 2S albumin, constitute 60-70 and 15-25% of total sesame proteins, respectively. Two gene families separately encoding four 11S globulin and three 2S albumin isoforms were identified in a database search of 3328 expressed sequence tag (EST) sequences from maturing sesame seeds. Full-length cDNA sequences derived from these two gene families were completed by PCR using a maturing sesame cDNA library as the template. The amino acid compositions of these deduced storage proteins revealed that the richness in methionine is attributed mainly to two 2S albumin isoforms and partly to one 11S globulin isoform. The presence of four 11S globulin and three 2S albumin isoforms resolved in SDS-PAGE was confirmed by MALDI-MS analyses. The abundance of these isoforms was in accord with the occurrence frequency of their EST sequences in the database. A comprehensive understanding of these storage proteins at the molecular level may also facilitate the identification of allergens in crude sesame products that have caused severe allergic reactions increasingly reported in the past decade.

Partition functions and finite-size scalings of Ising model on helical tori.

The exact closed forms of the partition functions of a two-dimensional Ising model on square lattices with twisted boundary conditions are given. The constructions of helical tori are unambiguously related to the twisted boundary conditions by virtue of the SL(2, Z) transforms. The numerical analyses on the deviations of the specific-heat peaks away from the bulk critical temperature reveal that the finite-size effect of herical tori is independent of the chirality.

Investigation on critical structural motifs of ligands for triggering glucocorticoid receptor nuclear migration through molecular docking simulations.

The glucocorticoid receptor (GR), a transcription factor regulating gene expression in a ligand-dependent fashion, is known for flexibility in adapting various ligands with their structures ranging from steroid to non-steroid. However, in our previous study, GR shows a stringent discrimination against a set of steroid ligands with highly similar structures for triggering its nuclear migration. In order to resolve this puzzle, we employed molecular docking simulations to investigate the origin of this structural discrimination. By analyzing the docking orientations and the related ligand-GR interaction patterns, we found that the hydrophilicity mismatch between the docking ligand and the GR ligand-binding site is the main cause combined with the steric hindrance and structural rigidness of these steroid ligands. Furthermore, we utilized this knowledge to rationalize how the structure-binding interaction of non-steroid ligands triggers GR nuclear migration with their structures available in Protein Data Bank.

Site specificity on OH α-H abstraction reaction for a zwitterionic ß-hairpin peptide in aqueous solution: a theoretical investigation.

Protein backbone oxidation was investigated by studying the α-H abstraction reaction in a ß-hairpin peptide, called Chignolin (PDB ID 1UAO), with density functional theory calculation at B3LYP/6-31G(d,p) without any constraint. In order to stabilize the zwitterionic form of Chignolin with the salt bridges, the effects of aqueous solution were implemented by using microsolvation combined with a conductor-like polarizable continuum model (CPCM). Comparison between three glycine residues located at three different sites in Chignolin was used to examine the possible site specificity of this backbone oxidation. To construct the reaction profile of these α-H abstraction reactions, the pre- and postreactive complexes along with their associated transition states were located and verified with the intrinsic reaction coordinate (IRC) method. The bond dissociation energy and reaction rates of these OH α-H abstraction reactions were calculated with transition state theory. The differences in this abstraction reaction between the neutral and zwitterionic forms of Chignolin were also compared. A molecular dynamics simulation was implemented to study the explicit solvation effect on the abstracted Chignolin structure. The range of the simulation time scale covers from femtoseconds to microseconds, i.e., from onset of the abstraction to the abstracted products reaching thermal equilibrium. Our results show that there are three kinds of site-specificity in this abstraction reaction. The reactivity and stability of the abstraction products and their abstraction modes are all dependent on the location where OH attacks. Furthermore, the free energy landscapes of these abstraction products are distinctively different. This may imply that the pathological disorders or diseases caused by this type of radicals are also dependent on the abstraction location.

Binding orientation and specificity of calmodulin to rat olfactory cyclic nucleotide-gated ion channel.

Calmodulin (CaM), the primary intracellular Ca(2+) receptor, regulates a large number of key enzymes and controls a wide spectrum of important biological responses. Recognition between CaM and its target sequence in rat olfactory cyclic nucleotide-gated ion channel (OLFp) was investigated by circular dichroism (CD), fluorescence, and NMR spectroscopy. Fluorescence data showed the OLFp tightly bound to CaM with a dissociation constant of 12 nM in a 1:1 stoichiometry. Far-UV CD data showed that approximately 60% of OLFp residues formed α-helical structures when associated with CaM. NMR data showed that most of the (15)N-(1)H HSQC cross-peaks of the (15)N-labeled CaM not only shifted but also split into two sets of peaks upon association with the OLFp. Our data indicated that the two distinct CaM/OLFp complexes existed simultaneously with stable structures that were not interexchangeable within the NMR time scale. In light of the palindromic sequence of OLFp (FQRIVRLVGVIRDW) for CaM targeting, we proposed that the helical OLFp with C2 symmetry may bind to CaM in two orientations. This hypothesis is supported by the observation that only one set of (15)N-(1)H HSQC cross-peaks of the (15)N-labeled CaM was detected upon association with OLFp-M13 chimeric peptide (OLFMp), a mutated OLFp lacking the palindromic feature. The binding specificity of OLFMp to CaM was restored when the palindromic feature was destroyed. Binding modes of CaM/OLFp and CaM/OLFMp simulated by molecular docking were in accord with their distinct patterns observed in HSQC spectra. Our studies suggest that the palindromic residues in OLFp are crucial for the orientation-specific recognition by CaM.

High-level expression, purification and production of the fungal immunomodulatory protein-gts in baculovirus-infected insect larva.

Fip-gts, a fungal immunomodulatory protein (Fip) isolated from Ganoderma tsugae (gts), has been reported to possess therapeutic effects in the treatment of cancer and autoimmune disease. To cost-effectively produce Fip-gts and bypass the bottleneck involved in its time-consuming purification from G. tsugae, in this study, we incorporated the SP(bbx) secretion signal into recombinant baculovirus for expressing glycosylated and bioactive rFip-gts in baculovirus-infected insect cells and Trichoplusia ni larva. This is the first study to employ the aerosol infecting T. ni larva with recombinant baculovirus for economical and high-level production of foreign proteins. In this study, one purification could yield 10 mg of rFip-gts protein merely from ∼100 infected T. ni larvae by aerosol inoculation, corresponding to 5 L (5 × 109 cells) of the infected Sf21 culture. In addition, the rFip-gts purified from T. ni larvae could induce the expression of interleukin-2 in murine splenocytes with an immunoresponsive level similar to that induced by LZ-8 (a known potent immunomodulatory protein purified from Ling zhi, Ganoderma lucidum). Thus, our results demonstrated that the larva-based baculovirus expression system can successfully express rFip-gts with the assembling capability required for maintaining immunomodulatory and anticancer activity. Our approach will open a new avenue for the production of rFip-gts and facilitate the immunoregulatory activity of rFip-gts available in the future.

Oxygen radical-mediated oxidation reactions of an alanine peptide motif - density functional theory and transition state theory study.

BACKGROUND: Oxygen-base (O-base) oxidation in protein backbone is important in the protein backbone fragmentation due to the attack from reactive oxygen species (ROS). In this study, an alanine peptide was used model system to investigate this O-base oxidation by employing density functional theory (DFT) calculations combining with continuum solvent model. Detailed reaction steps were analyzed along with their reaction rate constants. RESULTS: Most of the O-base oxidation reactions for this alanine peptide are exothermic except for the bond-breakage of the Cα-N bond to form hydroperoxy alanine radical. Among the reactions investigated in this study, the activated energy of OH α-H abstraction is the lowest one, while the generation of alkylperoxy peptide radical must overcome the highest energy barrier. The aqueous situation facilitates the oxidation reactions to generate hydroxyl alanine peptide derivatives except for the fragmentations of alkoxyl alanine peptide radical. The Cα-Cß bond of the alkoxyl alanine peptide radical is more labile than the peptide bond. CONCLUSION: the rate-determining step of oxidation in protein backbone is the generation of hydroperoxy peptide radical via the reaction of alkylperoxy peptide radical with HO2. The stabilities of alkylperoxy peptide radical and complex of alkylperoxy peptide radical with HO2 are crucial in this O-base oxidation reaction.

Inhibition of Na(+)/K(+) -ATPase by antcins, unique steroid-like compounds in Antrodia camphorate.

The inhibition of Na(+)/K(+) -ATPase by versatile steroid-like compounds contributes to the putative therapeutic effects of many Chinese medicinal cardiac products via the same molecular mechanism triggered by cardiac glycosides. Five major steroid-like compounds, antcin A, B, C, H, and K were isolated from Niuchangchih (Antrodia camphorata), a unique Taiwan mushroom, and all inhibited Na(+)/K(+) -ATPase. Antcin A exhibited significantly higher inhibitory potency than the other four antcins, though weaker than ginsenoside Rh2 . In contrast, cortisone (an analogous steroid with anti-inflammatory effects stronger than antcin A) showed no detectable inhibitory potency. Molecular modeling has shown that antcins bind to Na(+)/K(+) -ATPase with the steroidal skeleton structurally upside-down in comparison with ginsenoside Rh2 . The inhibitory potency of antcin A is attributed to steroidal hydrophobic interaction within the binding pocket and the formation of three hydrogen bonds between its carboxyl group and two cationic residues around the cavity entrance of Na(+)/K(+) -ATPase. The presence of an additional carbonyl or hydroxyl group at C7 of the other four antcins leads to severe repulsion in the hydrophobic pocket, and thus significantly reduces inhibitory potency. It is proposed that antcin A is a bi-functional compound that exerts anti-inflammatory effects and that enhances blood circulation via two different molecular mechanisms.