DNA glycosylases catalyze scission of the N-glycosylic bond linking a damaged base to the DNA sugar phosphate backbone. Some of these enzymes carry out a concomitant abasic (apyrimidinic/apurinic(AP)) lyase reaction at a rate approximately equal to that of the glycosylase step. As a generalization of the mechanism described for T4 endonuclease V, a repair glycosylase/AP lyase that is specific for ultraviolet light-induced cis-syn pyrimidine dimers, a hypothesis concerning the mechanism of these repair glycosylases has been proposed. This hypothesis describes the initial action of all DNA glycosylases as a nucleophilic attack at the sugar C-1' of the damaged base nucleoside, resulting in scission of the N-glycosylic bond. It is proposed that the enzymes that are only glycosylases differ in the chemical nature of the attacking nucleophile from the glycosylase/AP lyases. Those DNA glycosylases, which carry out the AP lyase reaction at a rate approximately equal to the glycosylase step, are proposed to use an amino group as the nucleophile, resulting in an imino enzyme-DNA intermediate. The simple glycosylases, lacking the concomitant AP lyase activity, are propose to use some nucleophile from the medium, e.g. an activated water molecule. This paper reports experimental tests of this hypothesis using five representative enzymes, and these data are consistent with this hypothesis.

A series of new resveratrol analogues were designed and synthesized and their inhibitory activities against aromatase were evaluated. The crystal structure of human aromatase (PDB 3eqm) was used to rationalize the mechanism of action of the aromatase inhibitor 32 (IC 50 0.59 µM) through docking, molecular mechanics energy minimization, and computer graphics molecular modeling, and the information was utilized to design several very potent inhibitors, including compounds 82 (IC 50 70 nM) and 84 (IC 50 36 nM). The aromatase inhibitory activities of these compounds are much more potent than that for the lead compound resveratrol, which has an IC 50 of 80 µM. In addition to aromatase inhibitory activity, compounds 32 and 44 also displayed potent QR2 inhibitory activity (IC 50 1.7 µM and 0.27 µM, respectively) and the high-resolution X-ray structures of QR2 in complex with these two compounds provide insight into their mechanism of QR2 inhibition. The aromatase and quinone reductase inhibitors resulting from these studies have potential value in the treatment and prevention of cancer.

ABSTRACT␤-N-Acetylhexosaminidases have attracted interest particularly for oligosaccharide synthesis, but their use remains limited by the rarity of enzyme sources, low efficiency, and relaxed regioselectivity of transglycosylation. In this work, genes of 13 ␤-Nacetylhexosaminidases, including 5 from Bacteroides fragilis ATCC 25285, 5 from Clostridium perfringens ATCC 13124, and 3 from Bifidobacterium bifidum JCM 1254, were cloned and heterogeneously expressed in Escherichia coli. The resulting recombinant enzymes were purified and screened for transglycosylation activity. A ␤-N-acetylhexosaminidase named BbhI, which belongs to glycoside hydrolase family 20 and was obtained from B. bifidum JCM 1254, possesses the bifunctional property of efficiently transferring both GalNAc and GlcNAc residues through ␤1-3 linkage to the Gal residue of lactose. The effects of initial substrate concentration, pH, temperature, and reaction time on transglycosylation activities of BbhI were studied in detail. With the use of 10 mM pNP-␤-GalNAc or 20 mM pNP-␤-GlcNAc as the donor and 400 mM lactose as the acceptor in phosphate buffer (pH 5.8), BbhI synthesized GalNAc␤1-3Gal␤1-4Glc and GlcNAc␤1-3Gal␤1-4Glc at maximal yields of 55.4% at 45°C and 4 h and 44.9% at 55°C and 1.5 h, respectively. The model docking of BbhI with lactose showed the possible molecular basis of strict regioselectivity of ␤1-3 linkage in ␤-N-acetylhexosaminyl lactose synthesis. IMPORTANCEOligosaccharides play a crucial role in many biological events and therefore are promising potential therapeutic agents. However, their use is limited because large-scale production of oligosaccharides is difficult. The chemical synthesis requires multiple protecting group manipulations to control the regio-and stereoselectivity of glycosidic bonds. In comparison, enzymatic synthesis can produce oligosaccharides in one step by using glycosyltransferases and glycosidases. Given the lower price of their glycosyl donor and their broader acceptor specificity, glycosidases are more advantageous than glycosyltransferases for large-scale synthesis. ␤-N-Acetylhexosaminidases have attracted interest particularly for ␤-N-acetylhexosaminyl oligosaccharide synthesis, but their application is affected by having few enzyme sources, low efficiency, and relaxed regioselectivity of transglycosylation. In this work, we describe a microbial ␤-N-acetylhexosaminidase that exhibited strong transglycosylation activity and strict regioselectivity for ␤-N-acetylhexosaminyl lactose synthesis and thus provides a powerful synthetic tool to obtain biologically important GalNAc␤1-3Lac and GlcNAc␤1-3Lac. Oligosaccharides are widely distributed in nature and play a crucial role in many biological events, such as cell structure modulation, cell-cell recognition and communication, and cellmicrobe/toxin interaction and adhesion; therefore, they are promising and important potential therapeutic agents (1-3). However, their use is limited because of the difficulty of large-scale production of oligosaccharides. Their...

Zephycandidine A (1), the first naturally occurring imidazo[1,2-f]phenanthridine alkaloid, was isolated from Zephyranthes candida (Amaryllidaceae). The structure of 1 was elucidated by spectroscopic analyses and NMR calculation, and a plausible biogenetic pathway for zephycandidine A (1) was proposed. Zephycandidine A (1) exhibited significant cytotoxicity against five cancer cell lines with IC50 values ranging from 1.98 to 7.03 μM with selectivity indices as high as 10 when compared to the normal Beas-2B cell. Further studies suggested that zephycandidine A (1) induces apoptosis in leukemia cells by the activation of caspase-3, upregulation of Bax, downregulation of Bcl-2, and degradation of PARP expression. In addition, zephycandidine A (1) showed acetylcholinesterase (AChE) inhibitory activity, and the docking studies of zephycandidine A (1) and galanthamine (2) with AChE revealed that interactions with W286 and Y337 are necessary.

Three pairs of new 8-O-4'-type dinorneolignan enantiomers, (±)-acortatarinowins A-C (1a/1b-3a/3b), a pair of new 8-O-4'-type (4a/4b) and a pair of rare C7-C8'-type (5a/5b) neolignan enantiomers, (±)-acortatarinowins D and E, and a pair of new furofuran-type lignan enantiomers, (±)-acortatarinowin F (6a/6b), along with two pairs of known lignan enantiomers (7a/7b and 8a/8b), were obtained from the rhizomes of Acorus tatarinowii. The separation of 1-8 by chiral HPLC using a Daicel IC column led to the isolation of eight pairs of enantiomers, 1a/1b-8a/8b, which had variable enantiomeric excess (ee) values of approximately 66, 71, 63, 60, 0, 38, 48, and 75%, respectively. The structures were elucidated by extensive spectroscopic and chemical methods, and their absolute configurations were determined by a combined analysis of single-crystal X-ray diffraction and a modified Mosher's method, assisted by experimental and calculated electronic circular dichroism data. Among them, compounds 1a, 3a, 6b, 8a, and 8b showed weak inhibitory activities against NO production in activated macrophages with IC50 values ranging from 23.3 to 38.0 μM, respectively.

Trastuzumab, the first targeted antibody against human epidermal growth factor receptor 2 (HER2), has been used to treat gastric cancer patients with HER2 overexpression. However, trastuzumab resistance often occurs following an initial period of benefits, and the underlying mechanisms remain largely unclear. The present study revealed that collagen type IV α1 chain (COL4A1), whose expression is upregulated in gastric cancer tissues and trastuzumab-resistant gastric cancer cells, may potentially confer trastuzumab resistance in gastric cancer. By performing bioinformatics analysis of 2 microarray datasets, the present study initially identified COL4A1, overexpressed in gastric cancer tissues and trastuzumab-resistant gastric cancer cells, as a potential candidate for inducing trastuzumab resistance. The drug resistance function of COL4A1 in gastric cancer was then validated by performing protein/gene interactions and biological process annotation analyses, and further validated by analyzing the functionality of microRNAs that target COL4A1 mRNA. Collectively, these data indicated that COL4A1 may confer trastuzumab resistance in gastric cancer.

Hyperisampsins A-D (1-4), with tetracyclo[6.3.1.1(3,10).0(3,7)]tridecane skeletons and seven biogenetically related congeners (5-11), were isolated from Hypericum sampsonii. Their structures were elucidated by comprehensive spectroscopic techniques. The absolute configuration of 1 was established by ECD calculations, and those of 5 and 9 were confirmed by single X-ray crystallographic analyses. Hyperisampsins A and D showed potent anti-HIV activities with EC50 of 2.97 and 0.97 μM and selectivity index of 4.80 and 7.70, respectively.

Ten new cytochalasan alkaloids, termed armochaetoglobins A-J (1-10), and four known chaetoglobosins (11-14) were isolated from a methanol extract of Chaetomium globosum TW1-1, a fungus isolated from the medicinal terrestrial arthropod Armadillidium vulgare. Their structures were elucidated by a combination of spectroscopy, single-crystal X-ray crystallography, and ECD calculations. Armochaetoglobins A-E (1-5) represented the first examples of seco-chaetoglobosins arising from an oxidative cleavage of C-19 and C-20. Among these compounds, armochaetoglobin A (1) features an unusual pyrrole ring. The cytotoxic activities of 2-10 were evaluated, and armochaetoglobin H (8) showed moderate inhibitory activities against five human cancer cell lines, with IC50 values ranging from 3.31 to 9.83 μM.