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C J Rizzo

Publications and source records attributed to C J Rizzo.

18 recordsLinked to original sources
PubMed2001

Stereocontrolled syntheses of all four stereoisomeric 1,N2-deoxyguanosine adducts of the lipid peroxidation product trans-4-hydroxynonenal.

[structure: see text]. trans-4-hydroxynonenal (4-HNE) is a unique product from the peroxidation of omega-6 polyunsaturated fatty acids. The major reaction of racemic 4-HNE with DNA is with deoxyguanosine to give four stereoisomeric exocyclic propano adducts. The stereospecific syntheses of these four adducts has been achieved at the nucleoside level. The synthetic approach is amenable to the synthesis of structurally defined oligonucleotides containing these endogenous genotoxins.

Aldehydes↗
PubMed2001

Absolute configuration and conformational stability of (+)-2,5-dimethylthiolane and (-)-2,5-dimethylsulfolane.

Enantiopure (+)-2,5-dimethylthiolane and (-)-2,5-dimethylsulfolane were prepared using literature procedures and investigated using vibrational circular dichroism (VCD). Experimental absorption and VCD spectra of (+)-2,5-dimethylthiolane and (-)-2,5-dimethylsulfolane in CCl(4) solution in the 2000-900 cm(-)(1) region were compared with the ab initio predictions of absorption and VCD spectra obtained with density functional theory using the B3LYP/6-311G(2d, 2p) basis set for different conformers of (2R,5R)-2,5-dimethylthiolane and (2R,5R)-2,5-dimethylsulfolane. This comparison indicates that (+)-2,5-dimethylthiolane is of the (2R,5R)-configuration and has two predominant conformations in CCl(4) solution. In addition, (-)-2,5-dimethylsulfolane is of (2R,5R)-configuration and has only one predominant conformation. The stereochemical assignment is in agreement with literature.

Journal Article↗
PubMed2001

Absolute configuration, predominant conformations, and vibrational circular dichroism spectra of enantiomers of n-butyl tert-butyl sulfoxide.

Alkylation of the alpha-carbanion of (R)-(-)-tert-butyl methyl sulfoxide (4) with n-propyl bromide afforded (+)-n-butyl tert-butyl sulfoxide (1) to which the absolute configuration (R) was ascribed. This assignment was confirmed by X-ray analysis of the complex 6 obtained from the enantiomerically pure sulfoxide (-)-1 and mercury chloride. Vibrational absorption and circular dichroism spectra of (+)-1 were measured in CDCl3 solution in the 2000-900 cm(-1) region and compared with the ab initio predictions of absorption and VCD spectra obtained with density functional theory using the B3LYP/6-31G basis set for different conformers of (R)-1. This comparison indicated also that (+)-1 is of the (R)-configuration.

Journal Article↗
PubMed2001

Synthesis of the C8-deoxyguanosine adduct of the food mutagen IQ.

[structure: see text] The C8-2'-deoxyguanosine adduct of the food mutagen 2-amino-3-methylimadazo[4,5-f]-quinoline (IQ) has been synthesized. The key step is a palladium-catalyzed N-arylation of a suitably protected 8-bromo-2'-deoxyguanosine derivative.

Biotransformation↗
PubMed2001

Inhibition of human endogenous retrovirus-K10 protease in cell-free and cell-based assays.

A full-length and C-terminally truncated version of human endogenous retrovirus (HERV)-K10 protease were expressed in Escherichia coli and purified to homogeneity. Both versions of the protease efficiently processed HERV-K10 Gag polyprotein substrate. HERV-K10 Gag was also cleaved by human immunodeficiency virus, type 1 (HIV-1) protease, although at different sites. To identify compounds that could inhibit protein processing dependent on the HERV-K10 protease, a series of cyclic ureas that had previously been shown to inhibit HIV-1 protease was tested. Several symmetric bisamides acted as very potent inhibitors of both the truncated and full-length form of HERV-K10 protease, in subnanomolar or nanomolar range, respectively. One of the cyclic ureas, SD146, can inhibit the processing of in vitro translated HERV-K10 Gag polyprotein substrate by HERV-K10 protease. In addition, in virus-like particles isolated from the teratocarcinoma cell line NCCIT, there is significant accumulation of Gag and Gag-Pol precursors upon treatment with SD146, suggesting the compound efficiently blocks HERV-K Gag processing in cells. This is the first report of an inhibitor able to block cell-associated processing of Gag polypeptides of an endogenous retrovirus.

Amino Acid Sequence↗
PubMed2001

Further computational studies on the conformation of 1,5-dihydrolumiflavin.

The optimized geometry of 1,5-dihydrolumiflavin has been calculated using density functional theory (DFT). Reduced lumiflavin was found to be bent along the N5-N10 axis, 25 degrees from planarity, which is nearly the same as previously reported restricted Hartree-Fock (RHF) calculations, which predict a bending angle of 27 degrees. The major difference in the DFT calculation is that the N10 methyl group adopts a more pseudoequatorial disposition and is only bent 13 degrees above the plane of the isoalloxazine ring system as opposed to 59 degrees in the RHF calculations. These computational results are compared with x-ray crystal structures of flavin models and flavoproteins. DFT calculation of 1,5-dihydroisoalloxazine resulted in a more modestly bent geometry of 17 degrees. This indicates that both electronic and steric interactions of the N10 methyl group of reduced lumiflavin contribute to the bent geometry.

Crystallography, X-Ray↗
PubMed2001

Inhibition of the bovine viral diarrhoea virus NS3 serine protease by a boron-modified peptidyl mimetic of its natural substrate.

Bovine viral diarrhoea virus (BVDV) is closely related to hepatitis C virus (HCV), and has been used as a surrogate virus in drug development for HCV infection. Similar to HCV, BVDV-encoded NS3 serine proteinase is responsible for multiple cleavages in the viral polyprotein, generating mature NS4A, NS4B, NS5A and NS5B proteins. NS3-dependent cleavage sites of BVDV contain a strictly conserved leucine at P1, and either serine or alanine at P1'. The full length BVDV NS3/4A serine protease has been cloned and expressed in bacterial cells. The enzyme has been purified from the soluble portion of Escherichia coli via a two-step purification procedure employing chromatography on heparin resin and gel filtration. The protease activity was characterized using in vitro translated BVDV NS4A/B and NS5A/B polyprotein substrates. A boronic acid analogue of the BVDV NS4A/NS4B cleavage site was synthesized and shown to be an efficient inhibitor of the NS3 serine protease in vitro. The compound, designated DPC-AB9144-00, inhibited approximately 75% of the NS3/4 activity at 10 microM with the NS4A/B substrate. However, no antiviral activity was detected with DPC-AB9144-00 in BVDV-infected Madin-Darby bovine kidney cells at concentrations as great as 90 pM, suggesting permeability or that other cellular-derived limitations were present.

Animals↗
PubMed2000

Stereocontrolled syntheses of deoxyribonucleosides via photoinduced electron-transfer deoxygenation of benzoyl-protected ribo- and arabinonucleosides.

The stereocontrolled, de novo syntheses of beta-2'-deoxy-, alpha-2'-deoxy-, beta-3'-deoxy-, and beta-2', 3'-dideoxyribonucleosides are described. Strategically protected ribose, arabinose, and xylose glycosylation precursors were synthesized bearing C2-esters capable of directing Vorbrüggen glycosylation. The key step is the regioselective deoxygenation of the desired hydroxyl group as either the benzoyl- or 3-(trifluoromethyl)benzoyl derivative. This deoxygenation is accomplished via a photoinduced electron-transfer (PET) mechanism using carbazole derivatives as the photosensitizer. The syntheses of the desired deoxynucleoside generally proceed in three steps from a common, readily available precursor.

Arabinonucleosides↗
PubMed1998

A cellular anti-apoptosis protein is cleaved by the HIV-1 protease.

Cleavage of non-viral proteins is rarely observed with the HIV-1 protease (HIV pr). One such cleavage event occurs with bcl-2, an important cytoprotective protein. The loss of bcl-2 has biological consequences, leading to enhanced HIV replication and programmed death of the host cell. A strategy is proposed to suppress HIV with non-cleavable mutants of bcl-2.

Apoptosis↗
PubMed1997

Molecular basis of HIV-1 protease drug resistance: structural analysis of mutant proteases complexed with cyclic urea inhibitors.

In cell cultures, the key residues associated with HIV-1 resistance to cyclic urea-based HIV-1 protease (PR) inhibitors are Val82 and Ile84 of HIV-1 PR. To gain an understanding of how these two residues modulate inhibitor binding, we have measured the Ki values of three recombinant mutant proteases, I84V, V82F, and V82F/I84V, for DMP323 and DMP450, and determined the three-dimensional structures of their complexes to 2.1-1.9 A resolution with R factors of 18.7-19.6%. The Ki values of these mutants increased by 25-, 0.5-, and 1000-fold compared to the wild-type values of 0.8 and 0.4 nM for DMP323 and DMP450, respectively. The wild-type and mutant complexes overall are very similar (rms deviations of 0.2-0.3 A) except for differences in the patterns of their van der Waals (vdw) interactions, which appear to modulate the Ki values of the mutants. The loss of the CD1 atom of Ile84, in the I84V mutant complexes, creates a hole in the S1 subsite, reducing the number of vdw contacts and increasing the Ki values. The V82F mutant binds DMP323 more tightly than wild type because the side chain of Phe82 forms additional vdw and edge-to-face interactions with the P1 group of DMP323. The Ki values of the single mutants are not additive because the side chain of Phe82 rotates out of the S1 subsite in the double mutant (the chi 1 angles of Phe82 and -182 in the V82F and V82F/I84V mutants differ by 90 and 185 degrees, respectively), further reducing the vdw interactions. Finally, compensatory shifts in the I84V and V82F/ I84V complexes pick up a small number of new contacts, but too few to offset the initial loss of interactions caused by the mutations. Therefore, our data suggest that variants persist in the presence of DMP323 and DMP450 because of a decrease in vdw interactions between the mutant proteases and inhibitors.

Azepines↗
PubMed1997

The HIV protease and therapies for AIDS.

New, potent therapies for HIV disease are available, based on synthetic inhibitors of the viral protease, an essential viral enzyme. The results in clinical trials have been impressive with most treated individuals benefiting in terms of reduced quantity of detectable virus, enhanced numbers of CD4 lymphocytes and improvements in quality and duration of life. However, there are some remaining negatives associated with the new drugs, including high cost, side effects and appearance of drug-resistant strains of HIV. Problems and future prospects for use of protease inhibitors and alternate approaches in AIDS are discussed.

Acquired Immunodeficiency Syndrome↗
PubMed1996

Apoptosis mediated by HIV protease is preceded by cleavage of Bcl-2.

Expression of the human immunodeficiency virus type 1 (HIV) protease in cultured cells leads to apoptosis, preceded by cleavage of bcl-2, a key negative regulator of cell death. In contrast, a high level of bcl-2 protects cells in vitro and in vivo from the viral protease and prevents cell death following HIV infection of human lymphocytes, while reducing the yields of viral structural proteins, infectivity, and tumor necrosis factor alpha. We present a model for HIV replication in which the viral protease depletes the infected cells of bcl-2, leading to oxidative stress-dependent activation of NF kappa B, a cellular factor required for HIV transcription, and ultimately to cell death. Purified bcl-2 is cleaved by HIV protease between phenylalanine 112 and alanine 113. The results suggest a new option for HIV gene therapy; bcl-2 muteins that have noncleavable alterations surrounding the HIV protease cleavage site.

Animals↗
PubMed1992

Molecular characterization of HIV-2 (ROD) protease following PCR cloning from virus infected H9 cells.

A 450 nucleotide sequence corresponding to the nucleotides 1931-2380 of the viral genome (8) was amplified by polymerase chain reaction (PCR) using template DNA prepared from HIV-2 (ROD) infected H9 cells. The sequence codes for HIV-2 protease and its N-terminal flanking peptide. An identical DNA sequence was obtained from three independent PCR amplifications, which differs from the published sequence of HIV-2 (ROD) in 7 nucleotides scattered throughout the region of the cloned DNA. The cloned DNA was expressed in E. coli cells and resulted in the synthesis of a correctly processed HIV-2 protease, which is enzymatically active. Therefore, none of the seven nucleotide changes, which resulted in two amino acid substitutions, affect the autoproteolytic or trans-cleaving activities of the HIV-2 protease.

Amino Acid Sequence↗
PubMed1991

An E. coli expression system which detoxifies the HIV protease.

Based on a variety of independent assays, the expression of HIV (human immunodeficiency virus type 1) protease in living bacterial cells results in their loss of viability. Although the mechanism is not proven, we have observed degradation of cellular proteins in E. coli expressing large amounts of active HIV protease. In order to avoid the loss of viability, we devised an expression system in which the viral protease is fused to beta-lactamase and is rapidly secreted to the periplasmic space, thus reducing its duration in the cytosol. Furthermore, we find the periplasmic form of the protease is soluble and enzymatically is several-fold more active than enzyme recovered from intracellular aggregates. The question of whether the viral protease may be toxic to infected cells is discussed.

Amino Acid Sequence↗