Cloning, overexpression, purification, and modeling of a lectin (Rhinocelectin) with antiproliferative activity from Tiger Milk Mushroom, Lignosus rhinocerus.

A lectin gene from the Tiger Milk Mushroom Lignosus rhinocerus TM02® was successfully cloned and expressed via vector pET28a in Escherichia coli BL21(DE3). The recombinant lectin, Rhinocelectin, with a predicted molecular mass of 22.8 kDa, was overexpressed in water-soluble form without signal peptide and purified via native affinity chromatography Ni-NTA agarose. Blast protein analysis indicated the lectin to be homologous to jacalin-related plant lectin. In its native form, Rhinocelectin exists as a homo-tetramer predicted with four chains of identical proteins consisting of 11 beta-sheet structures with only one alpha-helix structure. The antiproliferative activity of the Rhinocelectin against human cancer cell lines was concentration dependent and selective. The IC50 values against triple negative breast cancer cell lines MDA-MB-231 and breast cancer MCF-7 are 36.52 ± 13.55 µg mL-1 and 53.11 ± 22.30 µg mL-1 , respectively. Rhinocelectin is only mildly cytotoxic against the corresponding human nontumorigenic breast cell line 184B5 with IC50 value at 142.19 ± 36.34 µg mL-1 . The IC50 against human lung cancer cell line A549 cells is 46.14 ± 7.42 µg mL-1 while against nontumorigenic lung cell line NL20 is 41.33 ± 7.43 µg mL-1 . The standard anticancer drug, Doxorubicin exhibited IC50 values mostly below 1 µg mL-1 for the cell lines tested. Flow cytometry analysis showed the treated breast cancer cells were arrested at G0/G1 phase and apoptosis induced. Rhinocelectin agglutinated rat and rabbit erythrocytes at a minimal concentration of 3.125 µg mL-1 and 6.250 µg mL-1 , respectively.

The Antiproliferative Activity of Sclerotia of Lignosus rhinocerus (Tiger Milk Mushroom).

Lignosus rhinocerus, the tiger milk mushroom, is one of the most important medicinal mushrooms used by the indigenous people of Southeast Asia and China. It has been used to treat breast cancer. A cold water extract (LR-CW) prepared from the sclerotia of L. rhinocerus cultivar was found to exhibit antiproliferative activity against human breast carcinoma (MCF-7) and human lung carcinoma (A549), with IC(50) of 96.7 µg/mL and 466.7 µg/mL, respectively. In comparison, LR-CW did not show significant cytotoxicity against the two corresponding human normal cells, 184B5 (human breast cell) and NL 20 (human lung cell). DNA fragmentation studies suggested that the cytotoxic action of LR-CW against cancer cells is mediated by apoptosis. Sephadex G-50 gel filtration fractionation of LR-CW yielded a high-molecular-weight and a low-molecular-weight fraction. The high-molecular-weight fraction contains mainly carbohydrate (68.7%) and small amount of protein (3.6%), whereas the low-molecular-weight fraction contains 31% carbohydrate and was devoid of protein. Only the high-molecular-weight fraction exhibited antiproliferative activity against cancer cells, with IC(50) of 70.0 µg/mL and 76.7 µg/mL, respectively. Thus, the cytotoxic action of the LR-CW is due to the high-molecular-weight fraction, either the proteins or protein-carbohydrate complex.

Formation of colloidal suspension of hydrophobic compounds with an amphiphilic self-assembling peptide.

The amphiphilic self-assembling peptide EAK16-II was found to be able to stabilize hydrophobic compounds in aqueous solution. Micro/nanocrystals of a hydrophobic compound, pyrene, and a hydrophobic anticancer agent, ellipticine, were stabilized by EAK16-II to form colloidal suspensions in water. Initial evidence of the association between EAK16-II and hydrophobic compounds was the observation of a clouding phenomenon and a difference in fluorescence spectra of the solution. A further investigation on the interaction between EAK16-II and pyrene was carried out using fluorescence spectroscopy and scanning electron microscopy (SEM). It was found that the pyrene-peptide complex formation required mechanical stirring, and the freshly prepared peptide solution (containing peptide monomers and/or peptide protofibrils) was more effective at stabilizing pyrene than the mature fibrils in aged peptide solutions. The time duration over which the complex formed was about 22 h. The data on the complexation of pyrene and EAK16-II at various concentrations suggested that the maximum amount of stabilized pyrene was concentration dependent. SEM images showed that peptide concentration did not significantly affect the size of the complexes/suspensions but altered the structures of the peptide coating on the surface of the complex. Atomic force microscopy (AFM) was conducted to study the interaction of EAK16-II with a model hydrophobic surface, which provided some detailed information of how peptide adsorbed onto the hydrophobic compounds and stabilize them. This study shows the potential of self-assembling peptides for encapsulation of hydrophobic compounds.

Characterisation of a New Fungal Immunomodulatory Protein from Tiger Milk mushroom, Lignosus rhinocerotis.

Lignosus rhinocerotis (Tiger milk mushroom) is an important folk medicine for indigenous peoples in Southeast Asia. We previously reported its de novo assembled 34.3 Mb genome encoding a repertoire of proteins including a putative bioactive fungal immunomodulatory protein. Here we report the cDNA of this new member (FIP-Lrh) with a homology range of 54-64% to FIPs from other mushroom species, the closest is with FIP-glu (LZ-8) (64%) from Ganoderma lucidum. The FIP-Lrh of 112 amino acids (12.59 kDa) has a relatively hydrophobic N-terminal. Its predicted 3-dimensional model has identical folding patterns to FIP-fve and contains a partially conserved and more positively charged carbohydrates binding pocket. Docking predictions of FIP-Lrh on 14 glycans commonly found on cellular surfaces showed the best binding energy of -3.98 kcal/mol to N-acetylgalactosamine and N-acetylglucosamine. Overexpression of a 14.9 kDa soluble 6xHisFIP-Lrh was achieved in pET-28a(+)/BL21 and the purified recombinant protein was sequence verified by LC-MS/MS (QTOF) analysis. The ability to haemagglutinate both mouse and human blood at concentration ≥0.34 µM, further demonstrated its lectin nature. In addition, the cytotoxic effect of 6xHisFIP-Lrh on MCF-7, HeLa and A549 cancer cell lines was detected at IC50 of 0.34 µM, 0.58 µM and 0.60 µM, respectively.

Factors associated with breast self-examination behaviour among Chinese women in Hong Kong.

The purpose of this study was to examine factors predicting the practice of breast self-examination behaviour among Chinese women in Hong Kong using a cross-sectional survey research design. The Health Belief Model was used as the theoretical framework for the study to examine differences between breast self-examination practicers and non-practicers and among breast self-examination frequency groups. Data were collected from a convenience sample of 124 women using self-administrated questionnaires. Less than half of the sample practised breast self-examination and only 16% of the practicers performing breast self-examination monthly. There were differences between practicers vs. non-practicers and among breast self-examination frequency groups and some support of the predictive power of the Health Belief Model was found. Logistic regression showed that practicers perceived health as important, having fewer barriers and higher susceptibility to breast cancer. Discriminant function analysis revealed that barriers and children status were strong predictors of frequency of breast self-examination practice explaining 21.1% of the total variance in breast self-examination practice.

Practice of breast self-examination among high risk Chinese women in Hong Kong.

BACKGROUND: The incidence of breast cancer in Hong Kong is increasing and breast self-examination (BSE) as a screening tool is becoming more popular, especially among first-degree relatives (FDR) of breast cancer victims. BSE may be more applicable to Chinese women as their breast size is smaller. This study explores the BSE practice in this group of women. METHODS: A cross sectional study was conducted by sending questionnaires to 330 FDR of breast cancer patients treated in the Department of Surgery, University of Hong Kong. BSE behavior was studied with respect to its frequency, awareness, completeness and confidence of practice. RESULTS: 110 subjects returned the questionnaires with a response rate of 33%. The mean age of the respondent was 37 years. Only 57 women (52%) were practicers. The practicers have a stronger BSE awareness (P < 0.01) and a lower mean score on thought barriers (P = 0.002) than the non-practicers. 40% of the practicers gain their BSE knowledge through clinicians and their mean period of practice was 3.2 years. 68% of the practicers performed BSE completely. The overall confidence rate was 35%, but the rate was 43% among those who performed complete BSE. Thirteen factors possibly related to the completeness and rate of confidence of BSE examination were studied. The only factor that significantly determined completeness was the time spent for each examination (P = 0.002). The complete practicers required a longer time than the incomplete practicers (6.60 and 2.96 min, respectively). Women with a stronger BSE intention (P = 0.001) and a lower mean score on thought barriers (P = 0.001) performed the examination confidently. CONCLUSIONS: Slightly over half of the FDR practice BSE. The majority perform a complete BSE but they are not confident in finding abnormalities. Women spending more time on BSE are associated with a higher rate of completeness. However, only those with a stronger BSE intention and lesser thought barriers are more confident in their practice.

Protective effects of Mucuna pruriens seed extract pretreatment against cardiovascular and respiratory depressant effects of Calloselasma rhodostoma (Malayan pit viper) venom in rats.

The protective effects of Mucuna pruriens seed extract (MPE) against the cardio-respiratory depressant and neuromuscular paralytic effects induced by injection of Calloselasma rhodostoma (Malayan pit viper) venom in anaesthetized rats were investigated. While MPE pretreatment did not reverse the inhibitory effect of the venom on the gastrocnemius muscle excitability, it significantly attenuated the venom-induced cardio-respiratory depressant effects (p < 0.05). The protection effects may have an immunological mechanism, as indicated by the presence of several proteins in the venom that are immunoreactive against anti-MPE. However, we cannot rule out the possibility that the pretreatment may exert a direct, non-immunological protective action against the venom.

The protective effects of Mucuna pruriens seed extract against histopathological changes induced by Malayan cobra (Naja sputatrix) venom in rats.

Seed of Mucuna pruriens (Velvet beans) has been prescribed by traditional medicine practitioners in Nigeria as a prophylactic oral antisnake remedy. In the present studies, we investigated the protective effects of M. pruriens seed extract (MPE) against histopathological changes induced by intravenous injection of Naja sputatrix (Malayan cobra) venom in rats pretreated with the seed extract. Examination by light microscope revealed that the venom induced histopathological changes in heart and blood vessels in liver, but no effect on brain, lung, kidney and spleen. The induced changes were prevented by pretreatment of the rats with MPE. Our results suggest that MPE pretreatment protects rat heart and liver blood vessels against cobra venom-induced damages.

Solvent effect on the photophysical properties of the anticancer agent ellipticine.

This paper investigates how solution conditions, especially solvent polarity and hydrogen bonding, affect the fluorescence of ellipticine, a natural plant alkaloid with anticancer activity. A total of 16 solvents that cover a wide range of polarities were tested. The ultraviolet (UV) absorption and fluorescence emission of ellipticine were found to be solvent dependent. The absorption and emission maximum shifted to higher wavelengths (red shift) with increased solvent polarity. The difference in absorption and emission maximum (Stokes' shift) was large, approximately 10,000-11,000 cm-1, in polar solvents (with orientation polarizability Deltaf>0.2) but unusually small, approximately 8900 cm-1, in nonpolar solvents (hexane and cyclohexane). Large Stokes' shifts were due to an intramolecular charge transfer (ICT), which was enabled by large solvent polarity and hydrogen bonding of ellipticine with the solvents. Two transitions were found in the Lippert-Mataga plot between (1) nonpolar and semipolar solvents and between (2) semipolar and polar solvents. The first transition reflected the formation of hydrogen bonds between ellipticine and the solvents whereas the second transition indicated that ellipticine underwent an ICT. In addition, the larger extinction coefficients and the longer lifetime of ellipticine obtained in protic solvents were attributed to the formation of stronger hydrogen bonds. The photophysical response of ellipticine to changes in solvent polarity and hydrogen bond formation could be used to infer the location of ellipticine in a heterogeneous medium, namely liposomes in aqueous solution. A relatively large red shift of emission in liposomes indicated that ellipticine may be in a more polar environment with respect to the lipid bilayer, possibly close to the hydrophilic interface.

Biochemical and toxinological characterization of Naja sumatrana (Equatorial spitting cobra) venom

The lethal and enzymatic activities of venom from Naja sumatrana (Equatorial spitting cobra) were determined and compared to venoms from three other Southeast Asian cobras (Naja sputatrix, Naja siamensis and Naja kaouthia). All four venoms exhibited the common characteristic enzymatic activities of Asiatic cobra venoms: low protease, phosphodiesterase, alkaline phosphomonoesterase and L-amino acid oxidase activities, moderately high acetylcholinesterase and hyaluronidase activities and high phospholipase A2. Fractionation of N. sumatrana venom by Resource® S cation exchange chromatography (GE Healthcare, USA) yielded nine major protein peaks, with all except the acidic protein peak being lethal to mice. Most of the protein peaks exhibit enzymatic activities, and L-amino acid oxidase, alkaline phosphomonoesterase, acetylcholinesterase, 5'-nucleotidase and hyaluronidase exist in multiple forms. Comparison of the Resource® S chromatograms of the four cobra venoms clearly indicates that the protein composition of N. sumatrana venom is distinct from venoms of the other two spitting cobras, N. sputatrix (Javan spitting cobra) and N. siamensis (Indochinese spitting cobra). The results support the revised systematics of the Asiatic cobra based on multivariate analysis of morphological characters. The three spitting cobra venoms exhibit two common features: the presence of basic, potentially pharmacologically active phospholipases A2 and a high content of polypeptide cardiotoxin, suggesting that the pathophysiological actions of the three spitting cobra venoms may be similar.(AU)

Anion effect on the nanostructure of a metal ion binding self-assembling peptide.

Effects of copper salts containing different anions (SO(4)(2)(-), Cl(-), and NO(3)(-)) on the self-assembly of a designed peptide EAK16(II)GGH with affinity for Cu(2+) have been investigated. The peptide secondary structure, self-assembled nanostructures, and surface activity were observed to depend strongly on the type of anion. Over a salt concentration range from 0.05 to 10.0 mM, SO(4)(2)(-) induced long fiber formation, whereas Cl(-) and NO(3)(-) caused short fiber formation. The fiber length increased with copper sulfate concentration, but the concentration of copper chloride and copper nitrate did not affect the peptide nanostructures significantly. Analysis by Fourier transform infrared spectroscopy (FTIR) revealed that the addition of the copper salts tended to cause the peptide conformation to change from alpha-helix/random coil to beta-sheet, the extent to which depended on the anion type. This evidence of the anion effect was also supported by surface tension measurements using the axisymmetric drop shape analysis-profile (ADSA-P) technique. An explanation for the effect of anions on the peptide self-assembly was proposed. The divalent anion SO(4)(2)(-) might serve as a bridge by electrostatically interacting with two lysine residues from different peptide molecules, promoting beta-sheet formation. The extensive beta-sheet formation may further promote peptide self-assembly into long fibers. On the other hand, monovalent anions Cl(-) and NO(3)(-) may only electrostatically interact with one charged residue of the peptide; hence, a mixed secondary structure of alpha-helix/random coil and beta-sheet was observed. This observation might explain the predominant formation of short fibers in copper chloride and copper nitrate solutions.

Enzymatic and toxinological activities of Hypnale hypnale (hump-nosed pit viper) venom and its fractionation by ion exchange high performance liquid chromatography

Hypnale hypnale (hump-nosed pit viper) has been recently identified as one of the medically important venomous snakes in Sri Lanka and on the southwestern coast of India. The characterization of its venom is essential for understanding the pathophysiology of envenomation and for optimizing its management. In the present study, the biological properties of Hypnale hypnale venom and venom fractions obtained using Resource Q ion exchange chromatography were determined. The venom exhibited toxic activities typical of pit viper venom, comparable to that of its sister taxon, the Malayan pit viper (Calloselasma rhodostoma). Particularly noteworthy were its high activities of thrombin-like enzyme, proteases, phospholipase A2, L-amino acid oxidase and hyaluronidase. The thrombin-like enzyme was mainly acidic and distributed over several chromatography fractions, indicating its existence in multiple isoforms. The hemorrhagic and necrotic activities of the venom were likely associated with the proteolytic enzyme found mainly in the basic fraction. Phospholipase A2 and phosphomonoesterase exist in both acidic and basic isoforms, while L-amino acid oxidase and hyaluronidase are highly acidic. The venom clotting activity on fibrinogens showed distinct species specificity in the following increasing order for clotting time: bovine < rabbit < goat < human < horse < < dog, and was comparable to that of C. rhodostoma venom. Its clot formation on human fibrinogen is gradual and prolonged, a phenomenon suggestive of consumptive coagulopathy as a complication observed clinically. At an intramuscular sublethal dose, the venom did not cause acute kidney injury in a rodent model, contrary to the positive control group treated with Daboia russelii venom. Nephrotoxicity may result from higher venom doses in the context of coagulopathy, as a complication provoked by venom hematoxicity.(AU)

Enzymatic and immunological properties of Bungarus flaviceps (red-headed krait) venom

Bungarus flaviceps (red-headed krait) venom presents an intravenous LD50 of 0.32 ìg/g and exhibits enzymatic activities similar to other Bungarus toxins. ELISA cross-reactions between anti-Bungarus flaviceps and a variety of elapid and viperid venoms were observed in the current study. Double-sandwich ELISA was highly specific, since anti-B. flaviceps serum did not cross-react with any tested venom, indicating that this assay can be used for species diagnosis in B. flaviceps bites. In the indirect ELISA, anti-B. flaviceps serum cross-reacted moderately with three different Bungarus venoms (9-18 percent) and Notechis scutatus venom, but minimally with other elapid and viperid toxins. The results indicated that B. flaviceps venom shares common epitopes with other Bungarus species as well as with N. scutatus. The lethality of the B. flaviceps venom was neutralized effectively by antiserum prepared against B. candidus and B. flaviceps toxins and a commercial bivalent elapid antivenom prepared against B. multicinctus and Naja naja atra venoms, but was not neutralized by commercial antivenoms prepared against Thai cobra, king cobra and banded krait. These data also suggested that the major lethal toxins of B. flaviceps venom are similar to those found in B. multicinctus and B. candidus venoms.(AU)

Coumarins inPrunus mahaleb and its herbivore, the small ermine mothYponomeuta mahalebellus.

Larvae of the small ermine mothYponomeuta mahalebellus were reared on foliage ofPrunus mahaleb, a plant known to contain coumarins. Thin-layer chromatography and gas chromatography showed that coumarin, umbelliferone, and herniarin were present in leaves ofP. mahaleb and in pupae and adults ofY. mahalebellus. Overall concentrations of simple coumarins in the plant and insect were, respectively, 0.54% and 0.003-0.004% (dry weight). The possible role of coumarins in the chemical defense of both the plant and insect is discussed.

Cyanogenesis in Passifloraceae.

A novel cyclopentenoid cyanogenic bis-glycoside has been isolated from Passiflora capsularis L. Its structure has been tentatively determined as 4-boivinosyl tetraphyllin B. The name passicapsin is proposed for this compound. Linamarin has been isolated from Passiflora warmingii Mast. Extracts of Passiflora perfoliata L. yielded a (probably glycosidic) cyclopentenone dervative, which possibly originated from an unstable glycoside having a free alpha-hydroxynitrile group.

Redesigning the substrate specificity of an enzyme: isocitrate dehydrogenase.

Despite the structural similarities between isocitrate and isopropylmalate, isocitrate dehydrogenase (IDH) exhibits a strong preference for its natural substrate. Using a combination of rational and random mutagenesis, we have engineered IDH to use isopropylmalate as a substrate. Rationally designed mutations were based on comparison of IDH to a similar enzyme, isopropylmalate dehydrogenase (IPMDH). A chimeric enzyme that replaced an active site loop-helix motif with IPMDH sequences exhibited no activity toward isopropylmalate, and site-directed mutants that replaced IDH residues with their IPMDH equivalents only showed small improvements in k(cat). Random mutants targeted the IDH active site at positions 113 (substituted with glutamate), 115, and 116 (both randomized) and were screened for activity toward isopropylmalate. Six mutants were identified that exhibited up to an 8-fold improvement in k(cat) and increased the apparent binding affinity by as much as a factor of 80. In addition to the S113E mutation, five other mutants contained substitutions at positions 115 and/or 116. Most small hydrophobic substitutions at position 116 improved activity, possibly by generating space to accommodate the isopropyl group of isopropylmalate; however, substitution with serine yielded the most improvement in k(cat). Only two substitutions were identified at position 115, which suggests a more specific role for the wild-type asparagine residue in the utilization of isopropylmalate. Since interactions between neighboring residues in this region greatly influenced the effects of each other in unexpected ways, structural solutions were best identified in combinations, as allowed by random mutagenesis.

Concentration effect on the aggregation of a self-assembling oligopeptide.

Concentration is a key parameter in controlling the aggregation of self-assembling oligopeptides. By investigating the concentration effects, an aggregation mechanism of EAK16-II is proposed. Depending on the critical aggregation concentration (CAC) of EAK16-II, the oligopeptide aggregates into protofibrils through seeding and/or a nucleation process. Protofibrils then associate with each other to form fibrils. The CAC was found to be approximately 0.1 mg/ml by surface tension measurements. The nanostructures of aggregates were imaged and analyzed by atomic force microscopy. Globular and fibrillar aggregates were observed, and their dimensions were further quantified. To ensure that the aggregates were formed in bulk solution, light scattering (LS) measurements were conducted to monitor the fibril formation with time. The LS profile showed two different rates of aggregation depending on whether the peptide concentration was above or below the CAC. At high concentrations, the LS intensity increased strongly at early times. At low concentrations, the LS intensity increased only slightly. Our study provides information about the nature of the oligopeptide self-assembly, which is important to the understanding of the fibrillogenesis occurring in conformational diseases and to many biomedical engineering applications.

Serum kinetics of Calloselasma rhodostoma (Malayan pit viper) venom components in a rabbit

The serum kinetics of Calloselasma rhodostoma (Malayan pit viper) venom - specifically two of its components, the major hemorrhagin (rhodostoxin) and a thrombin-like enzyme - was examined in a rabbit by double-sandwich enzyme-linked immunosorbent assay (ELISA). The animal received intramuscularly a 1.0-mg/kg dose of C. rhodostoma venom. The venom level in serum peaked 12 hours after the injection, followed by a gradual decline and finally reached low rates 72 hours after administration. The serum kinetic profile of venom components, however, did not correspond to the profile of the whole C. rhodostoma venom. The serum levels of the C. rhodostoma thrombin-like enzyme increased slowly and peaked only 48 hours post-injection. Then both thrombin-like enzyme and rhodostoxin remained at relatively high levels 72 hours after administration. Data suggest that various venom components bind to tissue at the injection site with different affinities and that conjugated venom components were continuously released into circulation at different rates. The prolonged high serum levels of both thrombin-like enzyme and hemorrhagin are consistent with the clinical picture of prolonged clotting deficiency in severe cases of C. rhodostoma envenomation. Our results also suggest that since venom components are being released into and eliminated from the circulation at different rates, the "average composition" of the venom antigen in the circulation changes over time. This implies that data from ELISA quantification of antigen levels from serum venom employing "whole venom" as reagent must be interpreted with care.(AU)

Butenolides in small ermine moths,Yponomeuta spp. (Lepidoptera: Yponomeutidae), and spindle-tree,Euonymus europaeus (Celastraceae).

A new butenolide, isosiphonodin [3-hydroxymethyl-2(5H)-furanone], along with a trace of siphonodin [4-hydroxymethyl-2(5H)-furanone], was isolated from fifth-instar larvae of the small ermine mothYponomeuta cagnagellus. Leaves of its host plant spindle-tree,Euonymus europaeus, were found to contain the same two butenolides with siphonodin being present as the major compound. TLC showed that isosiphonodin was also present in larvae or pupae of six other small ermine moths which did not feed on spindle-tree. InY. cagnagellus butenolides might be plant derived, while isosiphonodin in the other investigated small ermine moths is probably synthesized by the insect. The possible role of butenolides in the chemical defense of small ermine moths is discussed.

Phlorisovalerophenone synthase, a novel polyketide synthase from hop (Humulus lupulus L.) cones.

Phlorisovalerophenone synthase (VPS), a novel aromatic polyketide synthase, was purified to homogeneity from 4.2 mg protein extract obtained from hop (Humulus lupulus L.) cone glandular hairs. The enzyme uses isovaleryl-CoA or isobutyryl-CoA and three molecules of malonyl-CoA to form phlorisovalerophenone or phlorisobutyrophenone, intermediates in the biosynthesis of the hop bitter acids (alpha- and beta-acids). VPS is an homodimeric enzyme, with subunits of 45 kDa. The pI of the enzyme was 6.1. Km values of 4 microm for isovaleryl-CoA, 10 microm for isobutyryl-CoA and 33 microm for malonyl-CoA, were found. The amino-acid sequences of two peptides, obtained by digestion of VPS, showed that the enzyme is highly homologous to plant chalcone synthases. The functional and structural relationship between VPS and other aromatic polyketide synthases is discussed.