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T C Bruice

Publications and source records attributed to T C Bruice.

At least 55 records · Page 3Linked to original sources
PubMed1997

Is strong hydrogen bonding in the transition state enough to account for the observed rate acceleration in a mutant of papain?

Nitriles are good inhibitors for the cysteine protease papain. However, a single amino acid mutation (Gln-19 --> Glu-19) in the active site makes the mutant enzyme a good catalyst for nitrile hydrolysis. A theoretical approach was used to examine the differential transition state stabilization in the papain mutant relative to the wild-type enzyme. Based on this study, we concluded that strong hydrogen bonding in the transition state is responsible for the observed rate enhancement of 4 x 10(5).

Binding Sites↗
PubMed1997

Targeting E2F1-DNA complexes with microgonotropen DNA binding agents.

Microgonotropen (MGT) DNA binding drugs, which consist of an A+T-selective DNA minor groove binding tripyrrole peptide and polyamine chains attached to a central pyrrole that extend drug contact into the DNA major groove, were found to be extraordinarily effective inhibitors of E2 factor 1 (E2F1) association with its DNA promoter element (5'-TTTCGCGCCAAA). The most active of these drugs, MGT-6a, was three orders of magnitude more effective than distamycin and inhibited complexes between E2F1 and the dihydrofolate reductase promoter by 50% at 0.00085 microM. A relationship was found between the measured equilibrium constants for binding of MGTs to the A+T region of d(GGCGA3T3GGC)/d(CCGCT3A3CCG) and their inhibition of complex formation between E2F1 and the DNA promoter element. A representative of the potent MGT inhibitors was significantly more active on inhibition of E2F1-DNA complex formation compared with disruption of a preexisting complex.

Animals↗
PubMed1997

A microgonotropen branched decaaza decabutylamine and its DNA and DNA/transcription factor interactions.

The central pyrrole of a site-selective DNA minor groove binding tripyrrole peptide 1 has been attached to a branched decaaza decabutylamine via a -(CH-2)3-NHCO-(CH2)-3 linker to provide the decaaza-microgonotropen (8). The decaaza decabutylamine moiety of 8 was designed to have a much greater affinity to the phosphodiester linkages of the backbone of DNA. Employing Hoechst 33258 (Ht) as a fluorescent titrant, the equilibrium constants for the binding for of 8 to the hexadecameric duplex d(GGCGCA3T3GGCGG)/d(CCGCCA3T3GCGCC) and to calf thymus DNA were determined. The log of the product of equilibrium constants (log Kl1Kl2) for 1:1 and 1:2 complexes formation at A3T3 is 17 (35 degrees C). Results of studies of the inhibition of the binding of several proteins to target DNA are discussed. Binding of the E2F1 transcription factor to its DNA target is 50% inhibited at approximately 2 nM concentration of 8.

Animals↗
PubMed1996

Mechanism of aldehyde oxidation catalyzed by horse liver alcohol dehydrogenase.

The mechanism of oxidation of benzaldehyde to benzoic acid catalyzed by horse liver alcohol dehydrogenase (HLADH) has been investigated using the HLADH structure at 2.1 A resolution with NAD+ and pentafluorobenzyl alcohol in the active site [Ramaswamy et al. (1994) Biochemistry 33,5230-5237]. Constructs for molecular dynamics (MD) investigations with HLADH were obtained by a best-fit superimposition of benzaldehyde or its hydrate on the pentafluorobenzyl alcohol bound to the active site Zn(II)ion. Equilibrium bond lengths, angles, and dihedral parameters for Zn(II) bonding residues His67, Cys46, and Cys174 were obtained from small-molecule X-ray crystal structures and an ab initio-derived parameterization of zinc in HLADH [Ryde, U. (1995) Proteins: Struct., Funct., Genet. 21,40-56]. Dynamic simulations in CHARMM were carried out on the following three constructs to 100 ps: (MD1) enzyme with NAD+, benzaldehyde, and zinc-ligated HO-in the active site; (MD2) enzyme with NAD+ and hydrated benzaldehyde monoanion bound to zinc via the pro-R oxygen, with a proton residing on the pro-S oxygen; and (MD3) enzyme with NAD+ and hydrated benzaldehyde monoanion bound to zinc via the pro-S oxygen, with a proton residing on the pro-R oxygen. Analyses were done of 800 sample conformations taken in the last 40 ps of dynamics. Structures from MD1 and MD3 were used to define the initial spatial arrangements of reactive functionalities for semiempirical PM3 calculations. Using PM3, model systems were calculated of ground states and some transition states for aldehyde hydration, hydride transfer, and subsequent proton shuttling. With benzaldehyde and zinc-bound hydroxide ion in the active site, the oxygen of Zn(II)-OH resided at a distance of 2.8-5.5 A from the aldehyde carbonyl carbon during the dynamics simulation. This may be compared to the PM3 transition state for attack of the Zn(II)-OH oxygen on the benzaldehyde carbonyl carbon, which has an O...C distance of 1.877 A. HLADH catalysis of the aldehyde hydration would require very little motion aside from that in the ground state. Two simulations of benzaldehyde hydrate ligated to zinc (MD2 and MD3) both showed close approach of the aldehyde hydrate hydrogen to NAD+C4, varying from 2.3 to 3.3 A, seemingly favorable for the hydride transfer reaction. The MD2 configuration does not allow proton shuttling. On the other hand, when the pro-S oxygen is ligated to zinc (MD3), the proton on the pro-R oxygen averages 2.09 A from the hydroxyl oxygen of Ser48 such that initiation of shuttling of protons via Ser48 to the ribose 2'-hydroxyl oxygen to the 3'-hydroxyl oxygen to His51 nitrogen is sterically favorable. PM3 calculations suggest that this proton shuttle represents a stepwise reaction which occurs subsequent to hydride transfer. The PM3 transition state for hydride transfer based on the MD3 configuration has the transferring hydride 1.476 A from C4 of NAD+ and 1.433 A from the aldehyde alpha-carbon.

Alcohol Dehydrogenase↗
PubMed1996

Molecular mechanics calculations of the riboacetal internucleotide linkage in double and triple helices.

Structures of Watson-Crick base paired 15-nucleobase oligomer strands in A-type or B-type conformation in which one strand [a strand of alternating nucleotide and riboacetal thymidine nucleoside (RT) units, RP] is DNA and the other is composed of alternating nucleotides and riboacetal nucleosides have been studied by molecular mechanics. Analogously, oligomer strands of RNA in place of DNA have been modeled. The calculations indicate that the RP strand is more stable when complexed in an A-type duplex relative to a B-type form and that this conformational preference is presumably due to the more uniform nature of the former. Nearly planar ribose rings were more commonly observed in the minimized structures of the B-type DNA.RP duplexes as compared with A-type duplexes, despite the fact that planar ribofuranose rings are known to be energetically unfavorable in oligonucleotides. Computed relative stabilities of all duplexes containing the RP strand suggest that such heteroduplexes are less stable than the corresponding double-stranded DNA and double-stranded RNA species. These findings are in agreement with experimental results which show, when equivalent sequences were compared, that a DNA.RNA control forms a more stable duplex than RP hound to a complementary single-stranded RNA strand. In contrast, molecular mechanics studies of complementary triple-helical (DNA)2.RP, (DNA)2.DNA, and (DNA)2.RNA structures indicate that the binding of RP as a Hoogsteen strand stabilizes the underlying duplex to a greater extent compared with native oligonucleotides. These calculations suggest that puckering of the ribose ring in the riboacetal linkage leads to a more favorable interaction with a complementary nucleic acid target than the proposed planar geometry and that this puckering may account for the enhanced binding of RP to a double-stranded target.

Acetals↗
PubMed1996

Design and synthesis of ribonucleic guanidine: a polycationic analog of RNA.

Replacement of the phosphodiester linkages of the polyanion RNA with guanidinium linkers (represented by g) provides the polycation ribonucleic guanidine (RNG). An anticipated structure for the triple-helical hybrid [r(Up)9U.r(Ag)9A.r(Up)9U] is presented. A basic strategy for the synthesis of RNG oligomers is described. Synthetic procedures are provided for tetrameric adenosyl RNG [r(Ag)3A].

Base Sequence↗
PubMed1996

A microgonotropen pentaaza pentabutylamine and its interactions with DNA.

The central pyrrole of a site-selective DNA minor groove binding tripyrrole peptide (1) has been attached to N-protected pentaazapentacosanoic acid (17) via a -(CH2)3-NHCO-(CH2)3- linker to provide 19, subsequent deprotection provided the pentaaza microgonotropen 4. The polyamine moiety of 4 reaches out of the minor groove and binds to the phosphate backbone of DNA. We find when employing Hoechst 33258 (Ht) as a fluorescent titrant to follow binding of 4 to the hexadecameric duplex d(GGCGCAAATTTGGCGG)/(CCGCCAAATTTGCGCC) and by 1H NMR titration of d(CGCAAATTTGCG)2 with 4 that the latter forms both 1:1 and 2:1 dsDNA complexes. Certain aspects of the structure of 4:d(CGCAAATTTGCG)2 complex derived via 1H NMR are discussed. The electrophoretic mobilities of phi X-174 DNA digested with HaeIII endonuclease restriction fragments complexed to 4 shows that the latter brings about a greater conformational change in the DNA fragments than observed previously with other microgonotropens.

DNA↗
PubMed1995

Binding studies of cationic thymidyl deoxyribonucleic guanidine to RNA homopolynucleotides.

Deoxyribonucleic guanidine is a potential antisense agent that is generated via the replacement of the negative phosphodiester linkages of DNA [--O--(PO2-)--O--] with positively-charged guanidinium (g) linkages [--NH--C(==NH2+)--NH--]. A pentameric thymidyl deoxyribonucleic guanidine molecule [d(Tg)4T-azido] has been shown to base pair specifically to poly(rA) with an unprecedented affinity. Both double and triple strands consisting of one and two equivalents of d(Tg)4T-azido paired with one equivalent of poly(rA) are indicated by thermal denaturation experiments. At an ionic strength of 0.22, the five bases of d(Tg)4T-azido are estimated to dissociate from a double helix with poly(rA) at > 100 degrees C! The effect of ionic strength on thermal denaturation is very pronounced, with stability greatest at low ionic strengths. The method of continuous variation indicates that there is an equilibrium complex with a molar ratio of d(Tg) to r(Ap) or d(Ap) of 2:1. Based on this evidence, models of the structures of d(Tg)9T-azido bound to r(Ap)9A are proposed.

Antisense Elements (Genetics)↗
PubMed1995

Synthesis of a thymidyl pentamer of deoxyribonucleic guanidine and binding studies with DNA homopolynucleotides.

Replacement of the phosphodiester linkages of the polyanion DNA with guanidine linkers provides the polycation deoxynucleic guanidine (DNG). The synthesis of pentameric thymidyl DNA is provided. This polycationic DNG species binds with unprecedented affinity and with base-pair specificity to negatively charged poly(dA) to provide both double and triple helices. The dramatic stability of these hybrid structures is shown by their denaturation temperatures (Tm). For example, the double helix of the pentameric thymidyl DNG and poly(dA) does not dissociate in boiling water (ionic strength = 0.12), whereas the Tm for pentameric thymidyl DNA associated with poly(dA) is approximately 13 degrees C (ionic strength = 0.12). The effect of ionic strength on Tm for DNG complexes with DNA shows an opposite correlation compared with double-stranded DNA and is much more dramatic than for double-stranded DNA.

Base Composition↗
PubMed1995

Electron tunneling and ab initio calculations related to the one-electron oxidation of NAD(P)H bound to catalase.

Models for NAD(P)H le- oxidation in bovine catalase were studied using Hartree-Fock ab initio calculations, along with information taken from the published X-ray structure of the enzyme. Geometries and energies of ground states and transition states were calculated at the 6-31G* level for N-methyl-1,4-dihydropyridine and N-methyl-1,4-dihydronicotinamide undergoing the pathway (i) le- oxidation to yield the radical cation, (ii) general-base-catalyzed (hydroxide and/or imidazole) deprotonation of the radical cation to yield the neutral radical, and (iii) le- oxidation of the neutral radical to the N-methylpyridinium or N-methylnicotinamide cation. Barrier heights for deprotonation of the radical cation intermediates were calculated to be 7-11 kcal/mol. Kinetic isotope effects were calculated for general-base-catalyzed deprotonation of the N-methyl-1,4-dihydropyridine radical cation and the 4,4-dideuterio species and found to be kH/kD2 = 5.38 (hydroxide) or 3.64 (imidazole), in qualitative agreement with published experimental isotope effects for the analogous deprotonation of N-methyl-1,10-dihydroacridan or the N-methyl-1,10-dideuterioacridan radical cation. In the calculated transition state for imidazole deprotonation of the N-methyl-1,4-dihydronicotinamide radical cation, an unusual short contact was calculated and interpreted as a hydrogen bond (2.35 A) between the amide oxygen and the hydrogen attached to C2 of imidazole. Similar hydrogen bonds were also observed and calculated at the 3-21G and 6-31G* levels between His234 of catalase and the amide oxygen of bound NAD(P)H and complexes of N,N'-dimethyl-1,4-dihydronicotinamide or cis-N-methylformamide with N-methylimidazole. Comparison of these results to the X-ray structure of bovine catalase allows for further interpretation of the possible roles of the imidazole bases His234 and His304 and the hydrogen-bonded contacts in the NAD(P)H binding site. Electron tunneling pathways between NAD(P)H and the iron protoporphyrin IX (PP-IX) axial tyrosinate ligand Tyr357 in molecular dynamics and X-ray crystal structures of bovine catalase were calculated using PATHWAYS II (version 2.01). The pathways which were calculated included those involving the amino acid residue Tyr214, which is near the NAD(P)H binding site. Coupling involving Tyr357 was not particularly efficient; however, strong coupling between Tyr214 and iron-protoporphyrin IX was observed. These pathways may be important if electron transfer is stepwise; i.e., Tyr214 oxidized first, followed by NAD(P)H.

Amino Acid Sequence↗
PubMed1995

NMR structure of d(CGCA3T3GCG)2:tren-microgonotropen-b:Zn(II) complex and solution studies of metal ion complexes of tren-microgonotropen-b interacting with DNA.

The solution structure of a 1:1 complex of zinc tren-microgonotropen-b [6b:Zn(II)] with d(CGCAAATTTGCG)2 has been determined by 2D nuclear Overhauser effect 1H NMR spectroscopy and restrained molecular modeling. The exchangeable and nonexchangeable proton resonances of d(CGCA3T3GCG)2:6b:Zn(II) indicate that the Zn(II) is interacting in the A+T-rich region of the dsDNA and the tren region of 6b, while 31P NMR shows interaction of the Zn(II) with the phosphate backbone. Proton chemical shift differences between d(CGCA3T3GCG)2:6b:Zn(II) and d(CGCA3T3GCG)2:6b are in agreement with the polyamino substituent of 6b [-(CH2)4N(CH2CH2)N-(CH2CH2NH2)2] forming a four-coordinated Zn(II) complex similar to that found in the X-ray structure of 'tren-chloride':Zn(II). The P9 and P10 phosphate oxygens that are held by hydrogen bonding to the tren substituent of 6b in the DNA:6b complex become ligands to the tren-complexed Zn(II) in DNA:6b:Zn(II). To do so there is a 2 A decrease in the adjacent phosphate-to-phosphate distance at the Zn(II) binding site. This motion brings about an increased bend of 14.6 degrees in the helical axis of d(CGCA3T3GCG)2:6b:Zn(II) compared to that found in d(CGCA3T3GCG)2:6b. Single stranded cleavage of linear DNA fragments was not observed in the presence of 6b and Fe(II), Co(II), Ni(II), Cu(II), Zn(II), La(III) or Ce(III); this is likely due to the metal ion being sequestered as in the structure of d(CGCA3T3GCG)2:6b:Zn(II) complex. Supercoiled DNA was susceptible to cleavage by 6b:Cu(II) in the presence of O2 and a reducing agent.

Base Sequence↗
PubMed1994

Design and synthesis of deoxynucleic guanidine: a polycation analogue of DNA.

The basic strategy is described for the connection of nucleosides by guanidinium (g) linkers to provide the positively charged deoxynucleic guanidine putative antigene agents. The synthetic procedures are provided for d(gT)n. Molecular modeling of double-stranded [d(gT)10.d(Ap)10] and the triple-helical hybrids [d(Tp)10.d(Ap)10.d(gT)10] and [d(gT)10.d(Ap)10.d(gT)10] suggest modes of interaction and anticipated structural features.

DNA↗
PubMed1994

Bending and straightening of DNA induced by the same ligand: characterization with the atomic force microscope.

A new ligand, MGT-6b, binds to DNA with two linked parts: a polyamine that binds to the phosphate backbone and a tripyrrole peptide that binds to the minor groove. This ligand decreases the curvature of bent kinetoplast DNA (kDNA) and also increases the curvature of a 400-bp DNA that is used as a molecular weight standard (M400), as characterized with the atomic force microscope (AFM). MGT-6b is more effective than distamycin at straightening kDNA. MGT-6b retards the electrophoretic mobility of M400 DNA, although neither of its two component parts alters the mobility of M400 or its curvature as seen in the AFM. Thus, both parts of the ligand are needed for the "vise grip" mode of binding that affects DNA bending. the ability of MGT-6b to both bend and straighten DNA is probably related to the very different DNA sequences and, hence, structures of kDNA and M400 DNA.

Bacteriophage phi X 174↗
PubMed1993

Stoichiometry and structure of complexes of DNA oligomers with microgonotropens and distamycin by 1H NMR spectroscopy and molecular modeling.

Dien-microgonotropen-c (5c), tren-microgonotropen-b (6b), and distamycin (Dm) bind the A.T-rich region of d(CGCAAATTTGCG)2 (oligo-12) and form 1:1 (5c and 6b) and 2:1 and 4:1 (Dm) complexes. At 1.75 mol ratio of Dm/oligo-12 the 4:1 complex starts to form and coexists with the 2:1 complex and the free double-stranded DNA. No 1:1 and 3:1 complexes were seen, implying a preferential dimeric binding mode of Dm to oligo-12. At 4:1 mol ratio of Dm/oligo-12 there is extensive exchange of the A.T imino protons with the solvent at the binding site. This is presumably due to the opening of the minor groove. Molecular modeling shows that four Dm molecules can fit in a tandem antiparallel way into the minor groove of oligo-12 by widening it to 16-17 A. On going from oligo-12 to the pseudosymmetrical hexadecamer oligo-16 [d(GGCGCAAATTTGGCGG).d(CCGCCAAATTTGCGCC)] the stoichiometry of binding of 5c changes from 1:1 to 2:1. Since oligo-12 and oligo-16 have the same A.T binding site this change in stoichiometry is due to the increase in the G.C terminal pairing. Hoechst 33258 displaces the two 5c molecules bound in the minor groove of oligo-16 at the A.T region. Marked exchange of A.T imino protons was seen in the case of (oligo-16).(Ht)2.

Base Composition↗
PubMed1993

Peptide nucleic acid (PNA) conformation and polymorphism in PNA-DNA and PNA-RNA hybrids.

Two hydrogen-bonding motifs have been proposed to account for the extraordinary stability of polyamide "peptide" nucleic acid (PNA) hybrids with nucleic acids. These interresidue- and intraresidue-hydrogen-bond motifs were investigated by molecular mechanics calculations. Energy-minimized structures of Watson-Crick base-paired decameric duplexes of PNA with A-, B-, and Z-DNA and A-RNA polymorphs indicate that the inherent stability of the complementary PNA helical structures is derived from interresidue, rather than from intraresidue, hydrogen bonds in all hybrids studied. Intraresidue-hydrogen-bond lengths are consistently longer than interresidue hydrogen bonds. Helical strand stability with interresidue hydrogen bond stabilization follows the order: B-(DNA.PNA) > A-(DNA.PNA) congruent to A-RNA.PNA > Z-(DNA.PNA). In the triplex hybrids A-(RNA.PNA2) and B-(DNA.PNA2), differences between stabilities of the two decamers of thyminyl PNA with lysine amide attached to the C terminus (pnaT)10 strands are small. The Hoogsteen (pnaT)10 strands are of slightly higher potential energy than are the Watson-Crick (pnaT)10 strands. Antiparallel arrangement of PNAs in the triplex is slightly favored over the parallel arrangement based on the calculations. Examination by molecular mechanics of the PNA.DNA analogue of the NMR-derived structure for the B-double-stranded DNA dodecamer d(CG-CAAATTTGCG)2 in solution suggests that use of all bases of the genetic alphabet should be possible without loss of the specific interresidue-hydrogen-bonding pattern within the PNA strand.

Base Sequence↗
PubMed1993

Molecular mechanics calculations of the structures of polyamide nucleic acid DNA duplexes and triple helical hybrids.

Polyamide nucleic acids (PNAs) have emerged as useful agents for recognition of single- and double-stranded nucleic acids. Interresidue hydrogen bonds between the amide carbonyl nearest the nucleobase and chain NH moieties provide inherent stability to the helical conformation of PNA 1. Moving the amide carbonyl away from the nucleobase to the backbone, and replacing it with a methylene group, results in 2 lacking the stabilizing hydrogen bond. Oligomers of 2 do not interact with DNA. Modeling suggests that 2 displays a more extended conformation than 1, and nucleobase orientation is disrupted in 2 in the absence of a complementary DNA strand. This is in contrast to 1, which retains a centrosymmetric arrangement of nucleobases. Structures for 1-T10.DNA and (1-T10)2.DNA species spanned by a pyrimidine strand (D-loop) were constructed. In the triple helical (1-T10)2.DNA structure, the two PNA strands form the complementary Watson-Crick paired strand and the Hoogsteen base-paired strand in the major groove of the 1.DNA duplex. The PNA strands are proposed to bind antiparallel to one another in (1-T10)2.DNA structure. The factors suggested to account for the stability of this 2:1 complex are (i) a hydrophobic attraction between two PNA backbones and (ii) a favorable electrostatic effect resulting from replacement of a phosphodiester backbone by a neutral peptide backbone.

Base Sequence↗
PubMed1992

Rational design of substituted tripyrrole peptides that complex with DNA by both selective minor-groove binding and electrostatic interaction with the phosphate backbone.

The structures of the compounds we call 3a, 3b, and 3c-compounds that incorporate (i) the tripyrrole peptide of the minor-groove-binding distamycin class of compounds and (ii) polyamine ligands that extend from the minor groove and can interact with phosphodiester bonds--were arrived at by computer-graphics designing by using the x-ray structure of distamycin A complexed in the minor groove of d(CGCAAATTTGCG)2. Compounds 3a, 3b, and 3c are elaborations of distamycin analog 2, designed for improved stability in solution and easier synthesis and purification, which itself binds weakly to DNA. Compounds 3a, 3b, and 3c have been synthesized, and the interaction of distamycin A, 2, 3a, 3b, and 3c with calf thymus DNA, poly(dA-dT), poly(dG-dC), poly(dI-dC), pBR322 superhelical plasmid DNA, and, in the case of 3b, T4 coliphage DNA have been studied. The following pertinent conclusions can be drawn. Binding of 3a, 3b, and 3c occurs in the minor groove of DNA and, because of favorable electrostatic interaction of diprotonated polyamine side chains and DNA phosphodiester linkages, the tenacity of DNA binding and site specificity of 3a, 3b, and 3c are comparable to that of native distamycin A. 3b has been found to induce changes in the superhelical density of pBR322 plasmid DNA. The study establishes that the central pyrrole N-CH3 substituent of 2 can be replaced by bulky polyamine metal ligands to create any number of compounds that bind into the minor groove at A + T-rich sites and are putative catalysts for the hydrolysis of DNA.

Animals↗