Synthesis, characterization and cytotoxicity of new platinum(IV) axial carboxylate complexes: crystal structure of potential antitumor agent [PtIV(trans-1R,2R-diaminocyclohexane)trans(acetate)2Cl2

A series of new platinum(IV) complexes of the type [PtIV(DACH)trans(L)2Cl2] (where DACH = trans-1R,2R-diaminocyclohexane, and L = acetate, propionate, butyrate, valerate, hexanoate, or heptanoate) bearing the carboxylate groups in the axial positions have been synthesized and characterized by elemental analysis, IR, and 195Pt NMR spectroscopy. The crystal structure of the analogue [PtIV(DACH)trans(acetate)2Cl2] was determined by single crystal X-ray diffraction method. There were two crystallographically independent molecules, both of which lie on crystallographic two-fold axes. The bond lengths and bond angles of both the molecules were the same within the experimental error. The compound crystallizes in the monoclinic space group C2, with a = 11.180(2) A, b = 14.736(3) A, c = 10.644(2) A, beta = 112.38(3) degrees, Z = 4 and R = 0.0336, based upon a total of 1648 collected reflections. In this complex, the platinum had a slightly distorted octahedron geometry owing to the presence of a geometrically strained five-member ring. The two adjacent corners of the platinum plane were occupied by the two amino nitrogens of DACH, whereas the other two equatorial positions were occupied by two chloride ions. The remaining two axial positions were occupied by the oxygens of acetate ligands. The DACH ring was in a chair configuration. An intricate network of intermolecular hydrogen bonds held the crystal lattice together. These analogues were evaluated in vitro and demonstrated cytotoxic activity against the human ovarian 2008 tumor cell line (IC50 = 0.001-0.06 microM). Structure-activity study revealed that activity was highest for the analogue where L = butyrate.     

Molecular Recombination Between R-Factor Deoxyribonucleic Acid Molecules in Escherichia coli Host Cells

THREE PREVIOUSLY STUDIED R FACTORS WERE USED: 222/R4, controlling transmissible resistance to sulfonamide, streptomycin, chloromycetin, and tetracycline (SU(r) SM(r) CM(r) TC(r)); 222/R3, a derivative of 222/R4 (now termed 222/R3W) having lost TC(r); and R15, controlling infectious resistance to SU and SM only. Two types of derivative R factors were isolated from 222/R4 by serial subculture in Salmonella species. One derivative, termed 222/R1, lost resistance to SU, SM, and CM, and the other, termed 222/R3N, lost only TC(r). Each factor was transferred to a standard Escherichia coli K-12 host. Recombinant factors of 222/R4 phenotype were isolated by selection after mixed culture of E. coli (222/R1)(+) and (222/R3N)(+) strains. Density-gradient equilibrium centrifugation of lysates of E. coli R(+) hosts in the presence of ethidium bromide separated R-factor deoxyribonucleic acid (DNA) as a heavy satellite peak which was subjected to electron microscopy or analytical density gradient centrifugation. Each DNA comprised a unimolecular species of circular DNA. The contour of R15 measured 22.3 mum [equivalent to 46 x 10(6) atomic mass units (AMU)], and that of 222/R4 measured 33.6 mum (70 x 10(6) AMU). 222/R3W appeared to be a point mutant or small deletion of 222/R4 with an almost identical size, whereas 222/R3N had lost a DNA segment of about 3 mum, and measured 30.3 mum or 63 x 10(6) AMU. The 222/R1 factors also appeared to have arisen by loss of DNA from 222/R4, 222/R1A being 22.3 mum or 46 x 10(6) AMU, whereas all other 222/R1 factors appeared to be duplicates, measuring 25.6 mum or 53 x 10(6) AMU. The DNA from six recombinant factors of R4 phenotype was indistinguishable in size and configuration from the parental 222/R4. In most cases, the number of R-factor copies (present as covalently closed circular molecules) per copy of the E. coli chromosome was less than 2, ranging from 1.2 to 3.3.     

Error-prone translesion synthesis by human DNA polymerase eta on DNA-containing deoxyadenosine adducts of 7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene

When human DNA polymerase eta (pol eta) encounters N6-deoxyadenosine adducts formed by trans epoxide ring opening of the 7,8-dihydroxy-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BaP DE) isomer with (+)-7R,8S,9S,10R configuration ((+)-BaP DE-2), misincorporation of A or G and incorporation of the correct T are equally likely to occur. On the other hand, the enzyme exhibits a 3-fold preference for correct T incorporation opposite adducts formed by trans ring opening of the (-)-(7S,8R,9R,10S)-DE-2 enantiomer. Adducts at dA formed by cis ring opening of these two BaP DE-2 isomers exhibit a 2-3-fold preference for A over T incorporation, with G intermediate between the two. Extension one nucleotide beyond these adducts is generally weaker than incorporation across from them, but among mismatches the (adducted A*) x A mispair is the most favored for extension. Because mutations can only occur if mispairs are extended, this observation is consistent with the occurrence of A x T to T x A transversions as common mutations in animal cells treated with BaP DE-2 isomers. Adducts with S absolute configuration at the point of attachment of the hydrocarbon to the base inhibit incorporation and extension by pol eta to a lesser extent than their R counterparts. Template-primers containing each of the four isomeric dA adducts derived from BaP DE-2 and two adducts derived from 9,10-epoxy-7,8,9,10-tetrahydrobenzo-[a]pyrene in which the 7- and 8-hydroxyl groups of the DEs are replaced with hydrogens exhibit reduced electrophoretic mobilities relative to the unadducted oligonucleotides. This effect is largely independent of DNA sequence. Decreased mobility correlates with an increased rate of incorporation by pol eta, suggesting a systematic relationship between the overall DNA structure and efficiency of the enzyme.     

Intercalation of the (1R,2S,3R,4S)-N6-[1-(1,2,3,4-tetrahydro-2,3,4-trihydroxybenz[a]anthracenyl)]-2'-deoxyadenosyl adduct in the N-ras codon 61 sequence: DNA sequence effects

The structure of the bay region (1R,2S,3R,4S)-N6-[1-(1,2,3,4-tetrahydro-2,3,4-trihydroxybenz[a]anthracenyl)]-2'-deoxyadenosyl adduct at X(7) of 5'-d(CGGACAXGAAG)-3'.5'-d(CTTCTTGTCCG)-3', incorporating codons 60, 61 (underlined), and 62 of the human N-ras protooncogene, was determined by NMR. This was the bay region benz[a]anthracene RSRS (61,3) adduct. The BA moiety intercalated above the 5'-face of the modified base pair. NOE connectivities between imino protons were disrupted at T16 and T17. Large chemical shifts at the lesion site were consistent with ring current shielding arising from the BA moiety. A large chemical shift dispersion was observed for the BA aromatic protons. An increased rise of 8.17 A was observed between base pairs A6 x T17 and X7 x T(16). The PAH moiety stacked with the purine ring of A6, the 5'-neighbor nucleotide. This resulted in buckling of the 5'-neighbor A6 x T17 base pair, evidenced by exchange broadening for the T17 imino resonance. It also interrupted sequential NOE connectivities between nucleotides C5 and A6. The A6 deoxyribose ring showed an increased percentage of the C3'-endo conformation. This differed from the bay region BA RSRS (61,2) adduct, in which the lesion was located at position X6 [Li, Z., Mao, H., Kim, H.-Y., Tamura, P. J., Harris, C. M., Harris, T. M., and Stone, M. P. (1999) Biochemistry 38, 2969-2981], but was similar to the benzo[a]pyrene BP SRSR (61,3) adduct [Zegar I. S., Chary, P., Jabil, R. J., Tamura, P. J., Johansen, T. N., Lloyd, R. S., Harris, C. M., Harris, T. M., and Stone, M. P. (1998) Biochemistry 37, 16516-16528]. The altered sugar pseudorotation at A6 appears to be common to both bay region BA RSRS (61,3) and BP SRSR (61,3) adducts. It could not be discerned if the C3'-endo conformation at A6 in the BA RSRS (61,3) adduct altered base pairing geometry at X7 x T16, as compared to the C2'-endo conformation. The structural studies suggest that the mutational spectrum of this adduct may be more complex than that of the BA RSRS (61,2) adduct.     

Specific molecular interaction between the insulin receptor and a D product of MHC class I

The density of MHC class I was determined on a murine thymoma cell line (R1), an H-2 negative variant (R1E), and R1E-derived cell lines in which H-2 expression was restored by transfection of various MHC class I genes (Db, Kb, and truncated Db) and/or a beta-2-microglobulin gene (beta 2-m; B2). Appreciable MHC class I expression was found on R1 cells and on the variants in which MHC class I expression was restored by transfection of Db/beta 2-m or Kb/beta 2-m genes. Only approximately 20% difference was observed between the number of Db molecules and Kb molecules on the R1E/B2/Db and on R1E/B2/Kb, respectively. However, specific insulin binding was significantly different between these lines. By using a computer assisted curve fitting program, the insulin binding data for R1 and R1E/B2/Db cell lines best fitted a two-site model (K approximately 6 x 10(-9) M for high-affinity sites and a 2 to 3 x 10(-7) M for low-affinity sites), whereas all other lines only expressed one type of insulin binding site. These sites were unrelated to IGF-I and IGF-II receptors. Cross-linking of 125I-labeled insulin demonstrated specific binding of the ligand to a Mr approximately 130,000 dalton band in all lines. In the R1E/B2/Db cells, insulin also cross-linked to cell membrane molecules with Mr approximately 48,000 and approximately 60,000 Da, which were identified by immunoprecipitation to be the H chain of MHC class I and the heavy chain of MHC class I plus beta 2-m, respectively. It is concluded that the insulin receptors in the cell membrane interact specifically with D-products of MHC class I and that class I molecules of MHC may have a crucial role in insulin receptor expression. This may reflect a more general nonimmunologic role of MHC class I.     

The rate constant describing slow-onset inhibition of yeast AMP deaminase by coformycin analogues is independent of inhibitor structure

(R)- and (S)-2'-deoxycoformycin, (R)-coformycin, and the corresponding 5'-monophosphates were compared as inhibitors of yeast AMP deaminase. The overall inhibition constants ranged from 4.2 mM for (S)-2'-deoxycoformycin to 10 pM for (R)-coformycin 5'-monophosphate, a difference of 3.8 x 10(8) in affinities. (R)-Coformycin, (R)-2'-deoxycoformycin 5'-monophosphate, and (R)-coformycin 5'-monophosphate exhibited both rapid and slow-onset inhibition. The S inhibitors and (R)-2'-deoxycoformycin exhibited classical competitive inhibition but no time-dependent onset of inhibition. The results indicate that the presence of the 2'-hydroxyl and 5'-phosphate and the R stereochemistry at the C-8 position of the diazepine ring are necessary for the optimum interaction of inhibitors with yeast AMP deaminase. This differs from the results for rabbit muscle AMP deaminase [Frieden C., Kurz, L. C., & Gilbert, H. R. (1980) Biochemistry 19, 5303-5309] and calf intestinal adenosine deaminase [Schramm, V. L., & Baker, D. C. (1985) Biochemistry 24, 641-646], in which a tetrahedral hydroxyl at C-8 in the R stereochemistry is sufficient for slow-onset inhibition with the coformycins. The results suggest that the transition state contains a tetrahedral carbon with the R configuration as a result of the direct attack of an oxygen nucleophile at C-6 of AMP. Slow-onset inhibition of yeast AMP deaminase is consistent with the mechanism [formula: see text] in which the combination of E and I is rapidly reversible. For these inhibitors, Ki varied by a factor of 3 x 10(3), and the overall inhibition constant (Ki*) varied by a factor of 2 x 10(5).(ABSTRACT TRUNCATED AT 250 WORDS)     

Palladium(II) salt and complexes of spermidine with a six-member chelate ring. Synthesis, characterization, and initial DNA-binding and antitumor studies.

By reaction of spermidine trihydrochloride with K2PdCl4 and PdCl2 at different pH's, we have synthesized the [sperH3]2[PdCl4]3 (I), [PdCl2(sperH)]2[PdCl4] (II), and [(PdCl2)3(sper)2] (III) compounds. The structure of these compounds was studied by IR and 1H NMR; complex II was analyzed by x-ray diffraction. In this complex the spermidine is attached to the PdCl2 group forming a six-member chelate ring with a protonated terminal amine group. The crystal of [PdCl2(sperH)]2[PdCl4] x 2H2O (II) is monoclinic, P2(1)/n, with a = 7.023(1) A, b = 12.662(1) A, c = 18.435(3) A, and beta = 99.95(1) degrees, Z = 4, R = 0.051, and Rw = 0.058 on the basis of 2690 independent reflections. We have compared the antitumor activity in vitro against the isolated human breast carcinoma MDA-MB 468 cell line of compounds I, II, and III with that of cis-diamminedichloroplatinum(II), cis-DDP. The results show that compounds III and III have values of ID50 similar (0.74 microgram/ml) or even lower (0.56 microgram/ml) than cis-DDP (0.80 microgram/ml). We also observed that compounds I, II, and III have the ability to induce conformational changes in covalently closed circular (ccc) form of the pUC8 plasmid DNA. Compounds II and III also induce conformational changes in the open circular (oc) form of this plasmid.     

Exploring QSAR of melatonin receptor ligand benzofuran derivatives using E-state index

Considering the recent challenge to the medicinal chemists for the development of selective melatonin receptor ligands, an attempt has been made to explore physicochemical requirements of benzofuran derivatives for binding with human MT1 and MT2 receptor subtypes and also to explore selectivity requirements. In this study, E-states of different common atoms of the molecules (calculated according to Kier and Hall) and physicochemical parameters (partition coefficient and molar refractivity) were used as independent variables along with suitable dummy parameters. The best equation describing MT1 binding affinity [n = 34, Q2 = 0.670, Ra2 = 0.790, R2 = 0.822, R = 0.907, s = 0.609, F = 25.8 (df 5, 28)] suggests that the binding affinity decreases as the value of n (number of CH2 spacer beside R2) increases while it increases with rise in electrotopological state values of different atoms of the benzofuran ring. Again, presence of methoxy group at R1 and hydrogen, unsubstituted phenyl or fluoro-substituted phenyl group at R2 is conducive to the MT1 binding affinity. The binding affinity decreases if furyl substitution at R3 position is present. The best equation describing MT2 binding affinity [n = 34, Q2 = 0.602, Ra2 = 0.755, R2 = 0.792, R = 0.890, s = 0.584, F = 213 (df 5, 28)] shows that the MT2 binding affinity depends on the similar factors as described for MT1 binding affinity; however, the contributions of the factors for the two affinities are different to some extent as evidenced from the regression coefficients. Among the selectivity relations, the best equation [n = 33, Q2 = 0.496 Ra2 = 0.681, R2 = 0.721, R = 0.849, s = 0.458, F = 18.1(df 4, 28)] suggests that MT2 binding increases with increase in value of n, presence of methoxy group at R1, and E-state values of different atoms of the benzofuran ring, while it decreases in presence of furyl group at R3 position.     

Probes for narcotic receptor mediated phenomena. Part 28: new opioid antagonists from enantiomeric analogues of 5-(3-hydroxyphenyl)-N-phenylethylmorphan

Enantiomeric analogues of 5-(3-hydroxyphenyl)morphan ligands were synthesized and evaluated because of our unexpected finding that opioid antagonists can be obtained in the 5-phenylmorphan series of opioids without sterically hindering the rotation of the phenolic ring. We determined the opioid receptor binding affinity of these new analogues, as well as the efficacy of the more interesting ligands. One of the new compounds [(1R,5S)-(-)-3-[2-(3'-phenylpropyl)-2-azabicyclo[3.3.1]non-5-yl]-phenol, 15] was found to have half of the efficacy of naloxone, a potent opioid antagonist, in the [(35)S]GTPgammaS assay, and two others (1R,5S)-(-)-3-[2-(4'-phenylbutyl)-2-azabicyclo[3.3.1]non-5-yl]-phenol, 17, and (1R,5S,1'S)-(+)-3-[2-(1'-methyl-2'-phenylethyl)-2-azabicyclo[3.3.1]non-5-yl]-phenol, 26, acted as moderately potent opioid antagonists. X-ray crystallographic structure data were obtained on three compounds. Two of them had three chiral centers; 25 [(1R,5S,1'R)-(-)-3-[2-(1'-methyl-2'-phenylethyl)-2-azabicyclo[3.3.1]non-5-yl]-phenol] was determined to have the 1R,5S,1'R configuration, and 26 the 1R,5S,1'S configuration. Since (1S,5R)-(+)-2-bromo-5-[2-(2'-phenylethyl)-2-azabicyclo[3.3.1]non-5-yl]-phenol (32) was a position isomer of (1S,5R)-(+)-4-bromo-3-[2-(2'-phenylethyl)-2-azabicyclo[3.3.1]non-5-yl]-phenol (30), and both showed the same 1H NMR spectrum, the structure of 32 was unequivocally determined by X-ray structure analysis.     

Intercalation of the (-)-(1R,2S,3R, 4S)-N6-[1-benz[a]anthracenyl]-2'-deoxyadenosyl adduct in an oligodeoxynucleotide containing the human N-ras codon 61 sequence

The solution structure of the (-)-(1R,2S,3R,4S)-N6-[1-(1,2,3, 4-tetrahydroxy-benz[a]anthracenyl)]-2'-deoxyadenosyl adduct at X6 of 5'-d(CGGACXAGAAG)-3'.5'-d(CTTCTTGTCCG)-3', incorporating codons 60, 61(italic), and 62 of the human N-ras protooncogene, was determined. This adduct results from the trans opening of 1S,2R,3R,4S-1, 2-epoxy-1,2,3,4-tetrahydro-benz[a]anthracenyl-3,4-diol by the exocyclic N6 of adenine. Molecular dynamics simulations were restrained by 509 NOEs from 1H NMR. The precision of the refined structures was monitored by pairwise root-mean-square deviations which were <1.2 A; accuracy was measured by complete relaxation matrix calculations, which yielded a sixth root R factor of 9.1 x 10(-)2 at 250 ms. The refined structure was a right-handed duplex, in which the benz[a]anthracene moiety intercalated from the major groove between C5.G18 and R,S,R,SA6.T17. In this orientation, the saturated ring of BA was oriented in the major groove of the duplex, with the aromatic rings inserted into the duplex such that the terminal ring of BA threaded the duplex and faced toward the minor groove direction. The duplex suffered localized distortion at and immediately adjacent to the adduct site, evidenced by the increased rise of 8.8 A as compared to the value of 3.5 A normally observed for B-DNA between base pairs C5.G18 and R,S,R,SA6.T17. These two base pairs also buckled in opposite directions away from the intercalated BA moiety. The refined structure was similar to the (-)-(7S,8R,9S,10R)-N6-[10-(7,8,9, 10)-tetrahydrobenzo[a]pyrenyl)]-2'-deoxyadenosyl adduct of corresponding stereochemistry at X6 of the same oligodeoxynucleotide [Zegar, I. S., Kim, S. J., Johansen, T. N., Horton, P. J., Harris, C. M., Harris, T. M., and Stone, M. P. (1996) Biochemistry 35, 6212-6224]. Both adducts intercalated toward the 5'-direction from the site of adduction. The similarities in solution structures were reflected in similar biological responses, when repair-deficient AB2480 Escherichia coli were transformed with M13mp7L2 DNA site-specifically modified with these two adducts.     

Role of a polycyclic aromatic hydrocarbon bay region ring in modulating DNA adduct structure: the non-bay region (8S,9R,10S, 11R)-N(6)-[11-(8,9,10,11-tetrahydro-8,9, 10-trihydroxybenz[a]anthracenyl)]-2'-deoxyadenosyl adduct in codon 61 of the human N-ras protooncogene.

The structure of the non-bay region (8S,9R,10S,11R)-N(6)-[11-(8,9,10, 11-tetrahydro-8,9,10-trihydroxybenz[a]anthracenyl)]-2'-de oxyadenosyl adduct at X(6) of 5'-d(CGGACXAGAAG)-3'.5'-d(CTTCTTGTCCG)-3', incorporating codons 60, 61 (underlined), and 62 of the human N-ras protooncogene, was determined. Molecular dynamics simulations were restrained by 475 NOEs from (1)H NMR. The benz[a]anthracene moiety intercalated above the 5'-face of the modified base pair and from the major groove. The duplex suffered distortion at and immediately adjacent to the adduct site. This was evidenced by the disruption of the Watson-Crick base pairing for X(6) x T(17) and A(7) x T(16) and the increased rise of 7.7 A between base pairs C(5) x G(18) and X(6) x T(17). Increased disorder was observed as excess line width of proton resonances near the lesion site. Comparison with the bay region benzo[a]pyrene [Zegar, I. S., Kim, S. J., Johansen, T. N., Horton, P. J., Harris, C. M., Harris, T. M., and Stone, M. P. (1996) Biochemistry 35, 6212-6224] and bay region benz[a]anthracene [Li, Z., Mao, H., Kim, H.-Y., Tamura, P. J., Harris, C. M., Harris, T. M., and Stone, M. P. (1999) Biochemistry 38, 2969-2981] adducts with the corresponding stereochemistry and at the same site shows that this non-bay region benz[a]anthracene lesion assumes different base pair geometry, in addition to exhibiting greater disorder. These differences are attributed to the loss of the bay region ring. The results suggest the bay region ring contributes to base stacking interactions at the lesion site. These structural differences between the non-bay and bay region lesions are correlated with site-specific mutagenesis data. The bay region benzo[a]pyrene and bay region benz[a]anthracene adducts were poorly replicated in vivo, and induced A --> G mutations. In contrast, the non-bay region benz[a]anthracene adduct was easily bypassed in vivo and was nonmutagenic.     

The stereochemistry of beta-lactam formation in cephalosporin biosynthesis.

3H and 14C from (2R,3S)[U-14C,3-3H1]cysteine and (2R,3R)-[U-14C,2,3-3H2]cysteine were incorporated into cephalosporin C by Cephalosporium acremonium. Analysis of the radioactive cephalosporin C indicated that the formation of its beta-lactam ring occurs stereospecifically and with retention of configuration at C-3 of cysteine.     

Molecular structure, stereochemistry and interactions of steffimycin B, and DNA binding anthracycline antibiotic

The crystal and molecular structure of anthracycline antibiotic steffimycin B(C29H32O13) has been determined by X-ray diffraction and the stereochemistry revealed. The orthorhombic crystals belong to space group P2(1)2(1)2(1), with the dimensions; a = 8.253 (2), b = 8.198 (2), c = 40.850 (8) A and Z = 4. Intensity data were collected for 2518 independent reflections. The structure was solved by direct methods and refined to an R value of 0.066 for 1410 reflections. The configuration in ring A is 7R,8S,9S. Ring A adopts half chair conformation, while the sugar ring has the regular chair conformation. The molecule most probably binds to double helical DNA through intercalation and hydrogen bonding.     

Synthesis and structure determination of some nonanomerically C-C-linked serine glycoconjugates structurally related to mannojirimycin

The Bucherer-Bergs reaction of methyl 2,3-O-isopropylidene-alpha-d-lyxo-hexofuranosid-5-ulose gave (4'S)-4'-carbamoyl-4'-[methyl (4R)-2,3-O-isopropylidene-beta-l-erythrofuranosid-4-C-yl]-oxazolidin-2'-one instead of expected hydantoins. A mixture of hydantoins--(5'R)-triphenylmethoxymethyl-5'-[methyl (4R)-2,3-O-isopropylidene-beta-l-erythrofuranosid-4-C-yl]-imidazolidin-2',4'-dione and (5'S)-triphenylmethoxymethyl-5'-[methyl (4R)-2,3-O-isopropylidene-beta-l-erythrofuranosid-4-C-yl]-imidazolidin-2',4'-dione was obtained from the 5-ulose having protected primary OH group at C-6. The 4'-S configuration of 2 as well as 5'-S configuration of (5'S)-hydroxymethyl-5'-[methyl (4R)-2,3-O-isopropylidene-beta-l-erythrofuranosid-4-C-yl]-imidazolidin-2',4'-dione (9) was confirmed by X-ray crystallography. Corresponding alpha-amino acid--methyl (5S)-5-amino-5-C-carboxy-5-deoxy-alpha-d-lyxo-hexofuranoside (alternative name: 2-[methyl (4R)-beta-l-erythrofuranosid-4-C-yl]-l-serine) (11) was obtained from the hydantoin 9 by acid hydrolysis of the isopropylidene and trityl groups followed by basic hydrolysis of the hydantoin ring. Analogous derivatives with 5-R configuration, formed in a minority, were also isolated and characterised.     

Variability in the backbone conformation of cyclic pentapeptides. Crystal structure of cyclic(Gly-L-Pro-D-Phe-Gly-L-Ala)

All the peptide bonds in cyclic(Gly-L-Pro-D-Phe-Gly-L-Ala) are in the trans conformation; however, the peptide bond C'5-N1 is twisted by 19 degrees from planarity (omega 5 = -161 degrees). A Type II beta-turn encompasses the L-Pro-D-Phe residues. Carbonyl oxygens O2, O4 and O5 are directed to the same side of the average plane through the backbone ring and they form hydrogen bonds with N3, N5 and N1, respectively, in adjacent molecules in a stacked column where the adjacent molecules are related by one translational unit. The conformation of the backbone is different from that established in other molecules with the DLDDL chirality sequence. The P21 cell contains two molecules of C21H26N5O5 with a = 4.836(2) A, b = 18.346(8) A, c = 12.464(5) A and beta = 100.05(4) degrees. The R factor for 1382 data with [F0[ greater than 1 sigma is 7.0%.     

Inhibition of class A and class C beta-lactamases by penems: crystallographic structures of a novel 1,4-thiazepine intermediate

A new beta-lactamase inhibitor, a methylidene penem having a 5,6-dihydro-8H-imidazo[2,1-c][1,4]oxazine heterocyclic substituent at the C6 position with a Z configuration, irreversibly inhibits both class A and class C serine beta-lactamases with IC(50) values of 0.4 and 9.0 nM for TEM-1 and SHV-1 (class A), respectively, and 4.8 nM in AmpC (class C) beta-lactamases. The compound also inhibits irreversibly the class C extended-spectrum GC1 beta-lactamase (IC(50) = 6.2 nM). High-resolution crystallographic structures of a reaction intermediate of (5R)-(6Z)-6-(5,6-dihydro-8H-imidazo[2,1-c][1,4]oxazin-2-ylmethylene)-7-oxo-4-thia-1-azabicyclo[3.2.0]hept-2-ene-3-carboxylic acid 1 with the SHV-1 beta-lactamase and with the GC1 beta-lactamase have been determined by X-ray diffraction to resolutions of 1.10 and 1.38 A, respectively. The two complexes were refined to crystallographic R-factors (R(free)) of 0.141 (0.186) and 0.138 (0.202), respectively. Cryoquenching of the reaction of 1 with each beta-lactamase crystal produced a common, covalently bound intermediate. After acylation of the serine, a nucleophilic attack by the departing thiolate on the C6' atom yielded a novel seven-membered 1,4-thiazepine ring having R stereochemistry at the new C7 moiety. The orientation of this ring in each complex differs by a 180 degrees rotation about the bond to the acylated serine. The acyl ester bond is stabilized to hydrolysis through resonance stabilization with the dihydrothiazepine ring and by low occupancy or disorder of hydrolytic water molecules. In the class A complex, the buried water molecule on the alpha-face of the ester bond appears to be loosely bound or absent. In the class C complex, a water molecule on the beta-face is disordered and poorly activated for hydrolysis. Here, the acyl intermediate is unable to assist its own hydrolysis, as is thought to occur with many class C substrates.     

Olfactory discrimination among sex pheromone stereoisomers: chirality recognition by pink hibiscus mealybug males

Our previous field studies suggested that the two chiral centers in the sex pheromone of pink hibiscus mealybug, Maconellicoccus hirsutus, could elicit different male responses. The chiral center in the acid moiety of the pheromone seemed to be more critical than the alcohol portion of the pheromone molecule for attractiveness. The objective of the current study was to test this hypothesis by deploying stereoisomeric blends in pheromone traps. Captures of male M. hirsutus showed that pheromone with the naturally occurring (R)-maconelliyl (S)-2-methylbutanoate and (R)-lavandulyl (S)-2-methylbutanoate [R-S configuration] was most attractive and that pheromone with the unnatural S-S configuration was less attractive. In addition, the RS-R blend (containing R-R and S-R stereoisomers) yielded captures of male M. hirsutus that were comparable to blank controls, and an inhibitory effect was observed when R-R and S-R were combined with naturally occurring R-S blend. These results suggest a unique chirality recognition mechanism; olfactory discrimination among different pheromone stereoisomers depends upon both asymmetric centers. The S configuration on the acid moiety elicits attraction, whereas the R configuration induces inhibition. However, the attractive activity shows some degree of tolerance toward chirality change in the alcohol portion of the pheromone molecules.     

Synthesis, antitubercular activity, and SAR study of N-substituted-phenylamino-5-methyl-1H-1,2,3-triazole-4-carbohydrazides

Tuberculosis treatment remains a challenge that requires new antitubercular agents due to the emergence of multidrug-resistant Mycobacterium strains. This paper describes the synthesis, the antitubercular activity and the theoretical analysis of N-substituted-phenylamino-5-methyl-1H-1,2,3-triazole-4-carbohydrazides (8a-b, 8e-f, 8i-j and 8n-o) and new analogues (8c-d, 8g-h, 8l-m and 8p-q). These derivatives were synthesized in good yields and some of them showed a promising antitubercular profile. Interestingly the N-acylhydrazone (NAH) 8n was the most potent against the Mycobacterium tuberculosis H37Rv strain (MIC=2.5 μg/mL) similar to or better than the current drugs on the market. The theoretical structure-activity relationship study suggested that the presence of the furyl ring and the electronegative group (NO(2)) as well as low lipophilicity and small volume group at R position are important structural features for the antitubercular profile of these molecules. NMR spectra, IR spectra and elemental analyses of these substances are reported.     

Biosynthesis of slaframine, (1S,6S,8aS)-1-acetoxy-6-aminooctahydroindolizine, a parasympathomimetic alkaloid of fungal origin. 3. Origin of the pyrrolidine ring.

The phytopathogen Rhizoctonia leguminicola has previously been shown to incorporate pipecolic acid into the piperidine alkaloids 1-acetoxy-6-aminooctahydroindolizine (slaframine) and 3,4,5-trihydroxyoctahydro-1-pyrindine. In the experiments described here, resting cultures of R. leguminicola were incubated with [1-14C]- and [2-14C]malonic acid and with [1-14C]- and [2-2H]acetic acid. Both acids were incorporated into the ring systems of both alkaloids. Mass spectrometric analysis of 2H-enriched slaframine showed that the label resides in the five-membered ring and that the methyl carbon of acetate is joined to the carboxyl carbon of pipecolate. A pipecolate-dependent decarboxylation of [1-14C]malonate was demonstrated in cell-free extracts of R. leguminicola. The results account for previously unattributed carbons in the two alkaloids and suggest the formation of an eight-carbon intermediate common to both alkaloids by acylation of malonate with pipecolic acid.