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Threats to several of the world's great animal migrations necessitate a research agenda focused on identifying drivers of their population dynamics. The monarch butterfly is an iconic species whose continental migratory population in eastern North America has been declining precipitously. Recent analyses have linked the monarch decline to reduced abundance of milkweed host plants in the USA caused by increased use of genetically modified herbicide-resistant crops. To identify the most sensitive stages in the monarch's annual multi-generational migration, and to test the milkweed limitation hypothesis, we analyzed 22 years of citizen science records from four monitoring programs across North America. We analyzed the relationships between butterfly population indices at successive stages of the annual migratory cycle to assess demographic connections and to address the roles of migrant population size versus temporal trends that reflect changes in habitat or resource quality. We find a sharp annual population decline in the first breeding generation in the southern USA, driven by the progressively smaller numbers of spring migrants from the overwintering grounds in Mexico. Monarch populations then build regionally during the summer generations. Contrary to the milkweed limitation hypothesis, we did not find statistically significant temporal trends in stage-to-stage population relationships in the mid-western or northeastern USA. In contrast, there are statistically significant negative temporal trends at the overwintering grounds in Mexico, suggesting that monarch success during the fall migration and re-establishment strongly contributes to the butterfly decline. Lack of milkweed, the only host plant for monarch butterfly caterpillars, is unlikely to be driving the monarch's population decline. Conservation efforts therefore require additional focus on the later phases in the monarch's annual migratory cycle. We hypothesize that lack of nectar sources, habitat fragmentation, continued degradation at the overwintering sites, or other threats to successful fall migration are critical limiting factors for declining monarchs.

The benzyl-ester bond linking the C-terminal delta-aminovaleric acid residue of a peptide to a polymeric support was cleaved with liquid hydrogen fluoride in the presence of anisole, added as scavenger. Instead of the expected peptide with a free carboxyl group at the C-terminus, a peptide terminating in a ketone derivative was obtained. The unusual extent of this known side-reaction was attributed to the effect of the distance between the amino group and the carboxyl group in the C-terminal residue. The results of model experiments corroborated this view.

Ahigh-throughput analytical characterization system was developed for quality control support of a central sample collection resource. This system utilizes liquid chromatography mass spectrometry with in-house developed data automation applications. Continuous operation of analytical instrumentation is accomplished by fully automating sample submission and report processing functions. Comprehensive analytical information characteristic of quality, chemical, and physical properties (e.g. relative purity, detection sensitivity, LogD) are automatically transferred to an on-line database. The application of this database for detailed quality assessment of a small sample library (ca. 24,000 compounds) is demonstrated.

This paper provides a detailed analysis of the various means available to US authorities for obtaining foreign evidence and other types of international assistance in money laundering cases. The means analysed here include mutual legal assistance treaties (MLATs) and similar processes; multilateral treaties; tax information exchange agreements (TIEAs) and tax treaties (for a narrow range of money laundering offences); court‐sponsored procedures for taking foreign depositions, including letters rogatory; the use of unilateral compulsory measures, such as subpoenas, for obtaining foreign evidence, and the use of FinCEN and Interpol resources. The initiatives of the G7, the Financial Action Task Force and the OECD regarding international cooperation in money laundering matters are also briefly treated.

The reactions of several alkynyl carbene complexes [(CO)5MC(OMe)CCR1, M = Cr, W, R1 = Me, Me3Si, Ph, i-Pr, t-Bu) with a variety of acyclic enol ethers and ketene acetals [CH2C(OR2)R3, R2 = Et, Me, i-Bu, SiMe2t-Bu, R3 = H, Me, EtO, p-MeC6H4] are examined. These reactions occur to give [2 + 2] cycloaddition products in all cases except with R1 = Me3Si where ene products predominate. The cyclobutenyl carbene complexes produced in the [2 + 2] cycloadditions undergo rapid electrocyclic ring-opening at room temperature when R3 = H to give butadienyl carbene complexes as the isolated products. The reactions of the alkynyl carbene complexes with cyclic enol ethers derived from cyclohexanone and cyclopentanone are more prone to give ene products than the acyclic enol ethers. Greater proportions of ene products are seen for six- rather than five-membered ring enol ethers and for silyl rather than alkyl enol ethers and for silyl rather than carbon substituents at R1. Only small differences are seen between chromium and tungsten complexes. The [2 + 2] cycloadditions with the E- and Z-isomers of ethyl prop-1-enyl ether are stereospecific with complexes in which R1 = Me but not with those with R1 = SiMe3. The cyclobutenyl carbene complexes from the latter reactions with tungsten derivatives were found to undergo stereoselective electrocyclic ring-opening at 70 °C to give only Z,E-butadienyl carbene complexes which result from the conrotatory ring-opening in which the ethoxy group rotates in an outward direction. An E,E-isomer was also isolated from the thermolysis mixture; however, it was shown not to be a primary product but rather the result of an isomerization of the Z,E-butadienyl carbene complex under the reaction conditions. The stereoselectivity of the electrocyclic ring-opening of these cyclobutenyl carbene complexes was shown to be the same as that found for their corresponding cyclobutenyl esters. In one case, an interesting cine-rearrangement of a cyclobutenyl carbene complex was observed. The metal can be oxidatively removed from the cyclobutenyl carbene complexes to give the corresponding cyclobut-1-enyl esters in good yield. Thus, alkynyl carbene complexes can serve as synthons for alkynyl esters in [2 + 2] cycloadditions with enol ethers and have the attractive feature of greatly increased reaction rates. Additional synthetic interest can be associated with processes in which the [2 + 2] cycloaddition of the alkynyl carbene complex is coupled in tandem with other reactions of the carbene complex functionality in the cycloadducts. This is illustrated with Diels–Alder reactions of the butadienyl carbene complexes and cyclohexadienone annulations of a cyclobutenyl carbene complex.

The proteolytic enzyme stromelysin-1 is a member of the family of matrix metalloproteinases and is believed to play a role in pathological conditions such as arthritis and tumor invasion. Stromelysin-1 is synthesized as a pro-enzyme that is activated by removal of an N-terminal prodomain. The active enzyme contains a catalytic domain and a C-terminal hemopexin domain believed to participate in macromolecular substrate recognition. We have determined the three-dimensional structures of both a C-truncated form of the proenzyme and an inhibited complex of the catalytic domain by X-ray diffraction analysis. The catalytic core is very similar in the two forms and is similar to the homologous domain in fibroblast and neutrophil collagenases, as well as to the stromelysin structure determined by NMR. The prodomain is a separate folding unit containing three alpha-helices and an extended peptide that lies in the active site of the enzyme. Surprisingly, the amino-to-carboxyl direction of this peptide chain is opposite to that adopted by the inhibitor and by previously reported inhibitors of collagenase. Comparison of the active site of stromelysin with that of thermolysin reveals that most of the residues proposed to play significant roles in the enzymatic mechanism of thermolysin have equivalents in stromelysin, but that three residues implicated in the catalytic mechanism of thermolysin are not represented in stromelysin.

The structures of ternary complexes of human inositol monophosphatase with inhibitory Gd3+ and either D- or L-myo-inositol 1-phosphate have been determined to 2.2-2.3 A resolution using X-ray crystallography. Substrate and metal are bound identically in each active site of the phosphatase dimer. The substrate is present at full occupancy, while the metal is present at only 35% occupancy, suggesting that Li+ from the crystallization solvent partially replaces Gd3+ upon substrate binding. The phosphate groups of both substrates interact with the phosphatase in the same manner with one phosphate oxygen bound to the octahedrally coordinated active site metal and another oxygen forming hydrogen bonds with the amide groups of residues 94 and 95. The active site orientations of the inositol rings of D- and L-myo-inositol 1-phosphate differ by rotation of nearly 60 degrees about the phosphate ester bond. Each substrate utilizes the same key residues (Asp 93, Ala 196, Glu 213, and Asp 220) to form the same number of hydrogen bonds with the enzyme. Mutagenesis experiments confirm the interaction of Glu 213 with the inositol ring and suggest that interactions with Ser 165 may develop during the transition state. The structural data suggest that the active site nucleophile is a metal-bound water that is activated by interaction with Glu 70 and Thr 95. Expulsion of the ester oxygen appears to be promoted by three aspartate residues acting together (90, 93, and 220), either to donate a proton to the leaving group or to form another metal binding site from which a second Mg2+ coordinates the leaving group during the transition state.

The structure of inositol monophosphatase has been determined to 2.60 A resolution in complexes with Mn2+ and with Mn2+ and phosphate. In the Mn2+ complex, three metal cations and one Cl were bound in the active site on each of the two subunits of the enzyme. Ligands to the three metals include the side chains of Glu 70, Asp 90, Asp 93, and Asp 220, t he carbonyl group of Ile 92, several solvent molecules and the chloride, which is a ligand to each of the cations. When phosphate is soaked into these Mn2+ cocrystals, one of the three Mn2+ ions is expelled from the active site, leaving metal ions with octahedral and tetrahedral coordination geometry. In addition, the structure of apoinositol monophosphatase was determined to 2.5 A resolution. Residues 70-75, a two-turn helical segment which is involved in metal coordination, moves away from the metal binding site by 2-3 A in the absence of cations. Residues 30-40, which wrap around the metal binding site and interact with the metal indirectly through solvent molecules and protein ligands to the metal, become disordered in the absence of metal. In various metal complexes, segmental mobility is also observed in the residues which form the metal binding sites. The results of these studies of the interaction of inositol monophosphatase with cations suggest that the enzyme accomplishes phosphate ester hydrolysis using two metal ions, one with octahedral and one with tetrahedral coordination geometry. Broad metal-binding specificity appears to result from extensive flexibility in several of the protein segments which contribute metal ligands, from the presence of alternate metal ligands and from metal coordination spheres which include water molecules.

The 3-dimensional structure of human carbonic anhydrase II (HCAII; EC 4.2.1.1) complexed with 3 structurally related inhibitors, 1a, 1b, and 1c, has been determined by X-ray crystallographic methods. The 3 inhibitors (1a = C8H12N2O4S3) vary only in the length of the substituent on the 4-amino group: 1a, proton; 1b, methyl; and 1c, ethyl. The binding constants (Ki's) for 1a, 1b, and 1c to HCAII are 1.52, 1.88, and 0.37 nM, respectively. These structures were solved to learn if any structural cause could be found for the difference in binding. In the complex with inhibitors 1a and 1b, electron density can be observed for His-64 and a bound water molecule in the native positions. When inhibitor 1c is bound, the side chain attached to the 4-amino group is positioned so that His-64 can only occupy the alternate position and the bound water is absent. While a variety of factors contribute to the observed binding constants, the major reason 1c binds tighter to HCAII than does 1a or 1b appears to be entropy: the increase in entropy when the bound water molecule is released contributes to the increase in binding and overcomes the small penalty for putting the His-64 side chain in a higher energy state.

A number of novel C-nucleosides related to purine derivatives are described in which the purine moiety has been replaced by the isosteric heterocycle, 8-aminoimidazo[1,2-α]pyrazine. The nucleosides prepared include the ribo, 3′-deoxy, 2′,3′-dideoxy, and 2′,3′-unsaturated derivatives. These C-nucleosides represent derivatives containing acid stable glycosyl bonds and they can be considered as analogs of adenine- or 3-deazaade-nine-containing nucleosides. Preparation of the parent ribonucleoside was accomplished by reaction of the C-l functionalized sugar, (2ξ)-1-amino-3,6-anhydro-l-deoxy-4,5-O-isopropylidene-7-O-trityl-D-allo-heptitol with 2,3-dichloropyrazine, followed by ring closure to the 8-chloroimidazo[1,2-α]pyrazine nucleoside, conversion to the 8-amino derivative and deblocking. A single crystal X-ray structure of the parent 8-amino-3-(β-D-ribofuranosyl)imidazo[1,2-α]pyrazine is described and the conformation compared to that of formycin. The sugar-modified analogs were prepared by subsequent functional group manipulations on the sugar moiety. Biological evaluation against HIV in H9 T-lymphoid cell culture showed the nucleosides to be devoid of significant antiviral activity compared to DDA. The 3-deazaadenosine analog also demonstrated weak suppression of mouse splenic NK activity toward YAC cells (mouse lymphoma cell targets). The imidazo[1,2-α]pyrazine analog of 3-deazaadenosine showed antiinflammatory activity in vivo in the rat pleurisy carrageenan model in the same range with 3-deazaadenosine.

L-685,818 differs only slightly in structure from the immunosuppressive drug FK-506, and both compounds bind with comparable affinity to the 12-kDa FK-506-binding protein (FKBP12), the major intracellular receptor for the drug. Despite these similarities, L-685,818 is a potent antagonist of both the immunosuppressive and toxic effects of the drug. Here, we present a structural analysis of this problem. Although FK-506 and L-685,818 differ greatly in pharmacology, we have found that the three-dimensional structures of their complexes with FKBP12 are essentially identical. Approximately half of each ligand is in contact with the receptor protein, and half is exposed to solvent; the exposed region includes the two sites where the compounds differ. These results indicate that the profound differences in the pharmacology of these two compounds are not caused by any difference in their interaction with FKBP12. Rather, these effects arise because relatively minor changes in the exposed part of a bound ligand have a strong effect on how FKBP12-ligand complexes interact with calcineurin, their putative intracellular target. In addition, FK-506 complexes with FKBP12 proteins from several species all inhibit mammalian calcineurin. Analysis of the three-dimensional structure of the complex with respect to residues conserved among these proteins suggests a small number of surface residues near the bound ligands that may play a critical role in interactions between the protein-drug complex and calcineurin.

The furan derivative (1) undergoes diastereoselective Diels-Alder reac- tions with dienophilic species such as α,β-unsaturated esters, nitriles, sulfones, and imides in good to excellent chemical yields. The oxobi- cyclic adducts obtained are amenable to chemical transformations which render them useful to natural product synthesis

The reactions of several alkynyl carbene complexes [(CO)5MC(OMe)CCR1, M = Cr, W, R1 = Me, Me3Si, Ph, i-Pr, t-Bu) with a variety of acyclic enol ethers and ketene acetals [CH2C(OR2)R3, R2 = Et, Me, i-Bu, SiMe2t-Bu, R3 = H, Me, EtO, p-MeC6H4] are examined. These reactions occur to give [2 + 2] cycloaddition products in all cases except with R1 = Me3Si where ene products predominate. The cyclobutenyl carbene complexes produced in the [2 + 2] cycloadditions undergo rapid electrocyclic ring-opening at room temperature when R3 = H to give butadienyl carbene complexes as the isolated products. The reactions of the alkynyl carbene complexes with cyclic enol ethers derived from cyclohexanone and cyclopentanone are more prone to give ene products than the acyclic enol ethers. Greater proportions of ene products are seen for six- rather than five-membered ring enol ethers and for silyl rather than alkyl enol ethers and for silyl rather than carbon substituents at R1. Only small differences are seen between chromium and tungsten complexes. The [2 + 2] cycloadditions with the E- and Z-isomers of ethyl prop-1-enyl ether are stereospecific with complexes in which R1 = Me but not with those with R1 = SiMe3. The cyclobutenyl carbene complexes from the latter reactions with tungsten derivatives were found to undergo stereoselective electrocyclic ring-opening at 70 °C to give only Z,E-butadienyl carbene complexes which result from the conrotatory ring-opening in which the ethoxy group rotates in an outward direction. An E,E-isomer was also isolated from the thermolysis mixture; however, it was shown not to be a primary product but rather the result of an isomerization of the Z,E-butadienyl carbene complex under the reaction conditions. The stereoselectivity of the electrocyclic ring-opening of these cyclobutenyl carbene complexes was shown to be the same as that found for their corresponding cyclobutenyl esters. In one case, an interesting cine-rearrangement of a cyclobutenyl carbene complex was observed. The metal can be oxidatively removed from the cyclobutenyl carbene complexes to give the corresponding cyclobut-1-enyl esters in good yield. Thus, alkynyl carbene complexes can serve as synthons for alkynyl esters in [2 + 2] cycloadditions with enol ethers and have the attractive feature of greatly increased reaction rates. Additional synthetic interest can be associated with processes in which the [2 + 2] cycloaddition of the alkynyl carbene complex is coupled in tandem with other reactions of the carbene complex functionality in the cycloadducts. This is illustrated with Diels–Alder reactions of the butadienyl carbene complexes and cyclohexadienone annulations of a cyclobutenyl carbene complex.

Inositol monophosphatase (EC 3.1.3.25), the putative molecular site of action of lithium therapy for manic-depressive illness, plays a key role in the phosphatidylinositol signaling pathway by catalyzing the hydrolysis of inositol monophosphates. To provide a structural basis from which to design better therapeutic agents for manic-depressive illness, the structure of human inositol monophosphatase has been determined to 2.1-A resolution by using x-ray crystallography. The enzyme exists as a dimer of identical subunits, each folded into a five-layered sandwich of three pairs of alpha-helices and two beta-sheets. Sulfate and an inhibitory lanthanide cation (Gd3+) are bound at identical sites on each subunit and establish the positions of the active sites. Each site is located in a large hydrophilic cavern that is at the base of the two central helices where several segments of secondary structure intersect. Comparison of the phosphatase aligned sequences of several diverse genes with the phosphatase structure suggests that the products of these genes and the phosphatase form a structural family with a conserved metal binding site.

A series of 5-substituted thieno[2,3-b]- and thieno[3,2-b)- and thieno[3,2-b)thiophene-2-sulfonamides was prepared and evaluated for topical ocular hypotensive activity in glaucoma models. The 5-substituents were varied to maximize both inhibitory potency against carbonic anhydrase and water solubility. At the same time, these substituents were varied in order to obtain compounds with the appropriate pKa to minimize pigment binding in the iris. All of these variables were optimized in the best compound, 5-[[(methoxyethyl)[(methoxyethyl)ethyl] amino]methyl]thieno[2,3-b]thiophene-2-sulfonamide hydrochloride (55).

The mode of binding of acetyl-pepstatin to the protease from the human immunodeficiency virus type 1 (HIV-1) has been determined by x-ray diffraction analysis. Crystals of an acetyl-pepstatin-HIV-1 protease complex were obtained in space group P2(1)2(1)2 (unit cell dimensions a = 58.39 A, b = 86.70 A, c = 46.27 A) by precipitation with sodium chloride. The structure was phased by molecular replacement methods, and a model for the structure was refined using diffraction data to 2.0 A resolution (R = 0.176 for 12901 reflections with I greater than sigma (I); deviation of bond distances from ideal values = 0.018 A; 172 solvent molecules included). The structure of the protein in the complex has been compared with the structure of the enzyme without the ligand. A core of 44 amino acids in each monomer, including residues in the active site and residues at the dimer interface, remains unchanged on binding of the inhibitor (root mean square deviation of alpha carbon positions = 0.39 A). The remaining 55 residues in each monomer undergo substantial rearrangement, with the most dramatic changes occurring at residues 44-57 (these residues comprise the so-called flaps of the enzyme). The flaps interact with one another and with the inhibitor so as to largely preserve the 2-fold symmetry of the protein. The inhibitor is bound in two approximately symmetric orientations. In both orientations the peptidyl backbone of the inhibitor is extended; a network of hydrogen bonds is formed between the inhibitor and the main body of the protein as well as between the inhibitor and the flaps. Hydrophobic side chains of residues in the body of the protein form partial binding sites for the side chains of the inhibitor; hydrophobic side chains of residues in the flaps complete these binding sites.

The reaction of 17 alpha-benzoyloxy-11 beta-hydroxy-3,20-dioxo-1, 4-pregnadien-21-al as the hemiacetal (1) with methanol:acetic acid:potassium cyanide:manganese dioxide followed by acetylation and preparative HPLC of the reaction mixture afforded 11 crystalline products. These products can be conveniently divided into three categories representing side-chain cleavage and oxidative esterification with or without elimination of the benzoyloxy group. Of special interest was the stereospecific formation of the C-17 cyanohydrin acetate 4a and the cis delta 17(20) enol acetate methyl ester 5. On the other hand, nonstereospecific addition of HCN to the side chain gave the C-20 epimeric cyanohydrin acetates 7a and 7b. The use of activated versus nonactivated MnO2 plays a major role in determining the quantitative distribution of the products. It was also discovered that even in the absence of MnO2, the reaction goes to completion. A proposed mechanism which explains the formation of all products is presented.

A series of carbohydrate-derived dienes with different patterns of substitution on the pyranose ring were synthesized and their Diels-Alder reactions investigated. The diene moiety was incorporated into the pyranose ring by oxidation of 4-O-methanesulfonate esters of sugar derivatives to enals, followed by Witting alkenation. This new class of dienes underwent cycloaddition with maleimide or its N-phenyl derivative to give annulated pyranosides. The Diels-Alder reactions were highly stereoselective, giving single products in some cases. Structural analysis of the reaction products was carried out by NMR spectroscopy and X-ray crystallography. The results indicated a strong preference for the formation of the products resulting from addition of the dienophile to the face of the diene opposite the anomeric center. In cases where the anomeric and allylic substituents on the diene occupied opposite faces, addition of the dienophile occurred predominantly from the face opposite the more remote anomeric center

An approach to rapidly process and interpret high-throughput liquid chromatography mass spectrometry data is presented. This approach applies an in-house developed computer application to process LC-MS report files containing spectral and chromatographic data from four different detectors (i.e. electrospray positive ionization, electrospray negative ionization mass spectrometry, UV absorption, and evaporative light scattering detection). Properties characteristic of detection and chromatographic retention are extracted and populated into a database. Approaches to applying this analytical information database for quality control analysis of ca. 400,000 samples are presented. Compound quality assessment methods employing average purity and detection data fields are compared to methods employing multiple quality control criteria (e.g. detection, purity, retention, and signal to noise). Structural similarity searches were applied with the analytical information database to identify compounds that may be undetectable by electrospray mass spectrometry. In addition, an approach to applying the database to aid in the selection of analytical detection and chromatography conditions for rapid analytical method development is also discussed.

The α- and β-forms of benzil monodimethylhydrazone have been shown to be the anti- and syn-isomers by X-ray crystallography.

The three-dimensional structure of the catalytic domain of stromelysin-1 complexed with an N-carboxyl alkyl inhibitor has been determined by NMR methods. The global fold consists of three helices, a five stranded beta-sheet and a methionine located in a turn near the catalytic histidines, classifying stromelysin-1 as a metzincin. Stromelysin-1 is unique in having two independent zinc binding sites: a catalytic site and a structural site. The inhibitor binds in an extended conformation. The S1' subsite is a deep hydrophobic pocket, whereas S2' appears shallow and S3' open.

Stromelysin-1, a member of the matrix metalloendoprotease family, is a zinc protease involved in the degradation of connective tissue in the extracellular matrix. As a step toward determining the structure of this protein, multidimensional heteronuclear NMR experiments have been applied to an inhibited truncated form of human stromelysin-1. Extensive 1H, 13C, and 15N sequential assignments have been obtained with a combination of three- and four-dimensional experiments. On the basis of sequential and short-range NOEs and 13C alpha chemical shifts, two helices have been delineated, spanning residues Asp-111 to Val-127 and Leu-195 to Ser-206. A third helix spanning residues Asp-238 to Gly-247 is characterized by sequential NOEs and 13C alpha chemical shifts, but not short-range NOEs. The lack of the latter NOEs suggests that this helix is either distorted or mobile. Similarly, sequential and interstrand NOEs and 13C alpha chemical shifts characterize a four-stranded beta-sheet with three parallel strands (Arg-100 to Ile-101, Ile-142 to Ala-147, Asp-177 to Asp-181) and one antiparallel strand (Ala-165 to Tyr-168). Two zinc sites have been identified in stromelysin [Salowe et al. (1992) Biochemistry 31, 4535-4540]. The NMR spectral properties, including chemical shift, pH dependence, and proton coupling of the imidazole nitrogens of six histidine residues (151, 166, 179, 201, 205, and 211), invariant in the matrix metalloendoprotease family, suggest that these residues are zinc ligands. NOE data indicate that these histidines form two clusters: one ligates the catalytic zinc (His-201, -205, and -211), and the other ligates a structural zinc (His-151, -166, and -179). Heteronuclear multiple quantum correlated spectra and specific labeling experiments indicate His-151, -179, -201, -205, and -211 are in the N delta 1H tautomer and His-166 is in the N epsilon 2H tautomer.

In the replication cycle of human immunodeficiency virus, viral mRNA is translated as a polyprotein precursor. Cleavage by a viral-encoded protease is required for release of mature viral structural proteins and enzymes from the precursor; the key role of the protease in the maturation of this virus, the causative agent in acquired immune deficiency syndrome (AIDS), makes the it an attractive target for therapeutic intervention in the treatment of AIDS. To aid in the design of safe and effective inhibitors, we have studied the structure of the protease from human immunodeficiency virus type 1 (HIV-1), both in its native state1 and in complex with a statine-containing inhibitor, acetyl-pepstatin2. We report here the determination of the structure of the protease in complex with a reduced-peptide inhibitor, L365,862 (Ser-Gln-Asn-PheΨ(CH2-N)Pro-Ile-Val-Gln). The amino acid sequence of this peptide corresponds to the cleavage site between the membrane-associated and capsid proteins in the polyprotein (with the substitution of Phe for Tyr); the replacement of the scissile bond in this octapeptide with a reduced-peptide linkage generates an inhibitor that has been used as an affinity ligand in the purification of the protease3. Although the structure of this complex is still being refined, a preliminary comparison to the structure of the complex between HIV-1 protease and acetyl-pepstatin reveals considerable difference in the length of hydrogen bonds between the protein and the inhibitor. The conformation of the backbone of most of the inhibitor is extended, but the backbone has a turn conformation at the position of the amino-terminal serine residue; turn-forming at this position may facilitate binding between the protease and its natural substrates.

The clinical efficacy of converting enzyme inhibitors1 for reducing blood pressure in a large percentage of hypertensive patients has aroused considerable interest in developing agents that interrupt the renin-angiotensin system at other points, for example by blockade of the angiotensin II receptor2 or by inhibition of the aspartic proteinase, renin. Substrate based design of renin inhibitors in which the scissile P1-P1′ dipeptide is replaced with a non-hydrolyzable group, often a mimetic of a tetrahedral transition state for amide bond hydrolysis, has provided a useful approach for obtaining a variety of inhibitor structure types,3 many with Ki’s of better than 10−9 M. A clinically useful renin inhibitor, however, has not yet emerged due to poor pharmacokinetics (i.e., metabolism or rapid clearance) or poor oral absorption, problems often encountered with peptidic drug targets. An example of this is seen with 1, a “tetrapeptide” inhibitor4 that spans the P4 through P3′ sites of the renin substrate and which utilizes the statine analog, ACHPA5, as a P1-P1′ dipeptide replacement (Figure 1). Although 1 is quite potent in vitro, very low levels of drug are found in the blood after oral administration to the rhesus monkey at 50 mg/kg, and the half life after intravenous administration is short (< 1 h). Rapid biliary excretion of intact drug has been demonstrated for a number of other renin inhibitors.6

The synthesis and biochemical evaluation of a series of oxadiazole derivatives of imidazobenzodiazepines related to the benzodiazepine antagonist Ro 15-1788 (2a) are reported. Although the oxadiazole ring is seen as an isosteric replacement for the ester linkage, significant differences in structure-activity trends were observed. Specifically, oxadiazoles 9-12 invariably had increased receptor efficacy (as witnessed by measurements of the GABA shift) relative to the corresponding ester. Additionally, and in direct contrast to the classical agonists such as diazepam, affinity for the benzodiazepine receptor was enhanced by a 7- rather than 8-halo substituent. The results are discussed in terms of a six-point receptor-binding model originally based on the X-ray structure of 2a. For comparison, the crystal structures of two representative oxadiazole derivatives, 10h and 12o, having a 6-oxo and 6-phenyl group, respectively, were determined and the data incorporated into a modified binding model to account for the greater efficacy of these compounds. It is concluded that the antagonist behavior of 2a relies upon the hydrogen-bond-acceptor properties of the ester carbonyl oxygen whereas for the oxadiazole series this site is localized at the imidazole nitrogen.

Emphysema is a chronic disease of the lower respiratory tract, characterized by the destruction of the alveolar walls of the lung (Robbins, et. al., 1984). It is a progressively degenerative disease for which there is no effective therapy at present. Human neutrophil elastase (HNE) is a serine protease which is stored in the azurophilic granules of neutrophils, and which has broad specificity for small aliphatic groups at the P1 position of substrates (Powers et. al., 1986). HNE digestion of elastin, the principal structural protein in the alveoli, is believed to be the molecular event that underlies the clinical manifestation of emphysema (Janoff, 1985). HNE is a single chain glycoprotein of 27,000 daltons molecular weight (Sinha et. al., 1987) which is synthesized during the development of the mature neutrophil (Clark et. al., 1984). Normally, proteolytic activity is released intracellularly within the neutrophil after the azurophilic granules entraining the enzyme are discharged into phagosomes containing engulfed debris (Parmley et. al., 1986). Tissue damage results when enzyme inevitably leaks out of the neutrophil (and into the extracellular space) by regurgitation during phagocytosis or by cell death, for example. To counter this dangerous condition, proteinaceous inhibitors have evolved that can rapidly inactivate HNE and arrest the process of tissue destruction. Some idea of the importance of these inhibitors is given by their concentration in serum, which is normally around 1.3 grams per liter for the alphas proteinase inhibitor (Carrell et. al.,1982). Health problems are noted when these concentrations drop below about 35% of normal (Carrell, 1986). Because alphal proteinase inhibitor is sensitive to oxidation, it has been suggested that the correlation between smoking and emphysema might be the result of an imbalance between free HNE and its inhibitors in the lung (Carp et. al., 1982).

The furan derivative (1) undergoes diastereoselective Diels-Alder reac- tions with dienophilic species such as α,β-unsaturated esters, nitriles, sulfones, and imides in good to excellent chemical yields. The oxobi- cyclic adducts obtained are amenable to chemical transformations which render them useful to natural product synthesis

The unsaturated aldehyde 1, a key intermediate in our approach to erythronolide A seco acid (2), has been prepared from the simple tetronic acid 6 by its seriatim transformations into the vinylogous urethanes 5,9, and 11 concluding with reductive conversion of the latter into 1

The crystal and molecular structures of paspaline and paspalicine, indolemevalonate metabolites from Claviceps paspali are presented.

A remakable reaction of the substituted bicyclo [3,3,0] oct-3-ene-2-one ( 2e ) with the sodium enolate of malonic ester is described.

The lithium enolates generated from the tetronic acid derived vinylogous urethanes 3 and 4 have been found to undergo alkylation reactions with a variety of electrophiles to give products with predictable stereochemistry accompanied by very high diastereoselectivity.

The regio- and stereochemistry of the intramolecular [2+2] photoproducts from two related vinylogous imides have been determined with the aid of X-ray crystallography; their chemistry and a mechanistic rationale of their formation are presented.

Synthese des composes du titre par oxydation des sulfoxydes et sulfones de phenothiazinol-3 et benzo [a] phenothiazinol-5

Human neutrophil elastase was inactivated by methoxysuccinyl-L-Ala-L-Ala-L-Pro-L-Ala-chloromethane. The modified enzyme was crystallized from 40 mM ammonium phosphate, pH 7.0 in the hexagonal space group P6(3) with unit cell parameters a = 74.53 A, b = 74.53 A, c = 70.88 A, alpha = beta = 90 degrees, gamma = 120 degrees. These crystals were resistant to radiation damage and diffracted beyond 1.84-A resolution. The asymmetric unit contained one 25,000-dalton monomer of human neutrophil elastase. Crystals were also grown from the enzyme modified with the analogous iodinated inactivator, p-iodoanilinosuccinyl-L-Ala-L-Ala-L-Pro-L-Ala-chloromethane. These crystals proved to be isomorphous with those of methoxysuccinyl-L-Ala-L-Ala-L-Pro-L-Ala-chloromethane-modified human neutrophil elastase, and served as a single-site, heavy atom derivative for solving the tertiary structure of the enzyme.

A new, potent hypocholesterolemic agent is produced by cultures of Aspergillus terreus. The isolation of the compound and its characterization as 4a, 5-dihydromevinolin containing a trans-fused octahydro-naphthalene system are described. Comparative data for dihydromevinolin and mevinolin in three biological assays are given: in vitro inhibition of HMG-CoA reductase, inhibition of sterol synthesis in cell cultures, and inhibition of cholesterol synthesis in vivo in rats.