Pyrimidine Portion Research Articles

Constrained NBMPR analogue synthesis, pharmacophore mapping and 3D-QSAR modeling of equilibrative nucleoside transporter 1 (ENT1) inhibitory activity

Conformationally constrained analogue synthesis was undertaken to aid in pharmacophore mapping and 3D-QSAR analysis of nitrobenzylmercaptopurine riboside (NBMPR) congeners as equilibriative nucleoside transporter 1 (ENT1) inhibitors. In our previous study [ J. Med. Chem. 2003, 46, 831–837], novel regioisomeric nitro-1,2,3,4-tetrahydroisoquinoline conformationally constrained analogues of NBMPR were synthesized and evaluated as ENT1 ligands. 7-NO 2-1,2,3,4-Tetrahydroisoquino-2-yl purine riboside was identified as the analogue with the nitro group in the best orientation at the NBMPR binding site of ENT1. In the present study, further conformational constraining was introduced by synthesizing 5′- O,8-cyclo derivatives. The flow cytometrically determined binding affinities indicated that the additional 5′- O,8-cyclo constraining was unfavorable for binding to the ENT1 transporter. The structure–activity relationship (SAR) acquired was applied to pharmacophore mapping using the PHASE program. The best pharmacophore hypothesis obtained embodied an anti-conformation with three hydrogen-bond acceptors, one hydrophobic center, and two aromatic rings involving the 3′-OH, 4′-oxygen, the NO 2 group, the benzyl phenyl and the imidazole and pyrimidine portions of the purine ring, respectively. A PHASE 3D-QSAR model derived with this pharmacophore yielded an r 2 of 0.916 for four ( 4) PLS components, and an excellent external test set predictive r 2 of 0.78 for 39 compounds. This pharmacophore was used for molecular alignment in a comparative molecular field analysis (CoMFA) 3D-QSAR study that also afforded a predictive model with external test set validation predictive r 2 of 0.73. Thus, although limited, this study suggests that the bioactive conformation for NBMPR at the ENT1 transporter could be anti. The study has also suggested an ENT1 inhibitory pharmacophore, and established a predictive CoMFA 3D-QSAR model that might be useful for novel ENT1 inhibitor discovery and optimization.

Unusual effect of bulky isopropyl group on robustness of the U-motif in pyrazolo[3,4- d]pyrimidine core based ‘Leonard linker’ compounds in comparison to methyl/ethyl group: A 1H NMR and X-ray crystallographic study

X-ray crystallographic analysis of new symmetrical ‘Leonard/trimethylene linker’ isopropyl analog 1,3-bis(4-isopropoxy-6-methylsulfonyl-1 H-pyrazolo[3,4- d]pyrimidin-1-yl)propane ( 1h) does not show chain motif formed due to intermolecular C H…O interactions between O atoms of sulfonyl groups and central methylene H atoms of linker along with C H…N interactions or plus (+) motif shown earlier by corresponding methyl- ( 1f) and ethyl- ( 1g) analogs. More importantly, the more or less superimposed U-motif conformation of 1h is very different from earlier related 10 symmetrical compounds ( 1a– 1g & 2). In 1h intramolecular folding is in such a way that whole pyrazolo[3,4- d]pyrimidine core of one side stacks more or less face-to-face on opposite side of the same molecule while in earlier compounds ( 1a– 1g & 2) there is no intramolecular stacking between pyrazolo portions and only pyrimidine portions partially stack with each other. Surprisingly, ethyl analog ( 1i) of 1h again shows U-motif conformation similar to earlier related ten symmetrical compounds ( 1a– 1g & 2). Supramolecular structures of both compounds ( 1h & 1i) are stabilized by weak intermolecular C H…O, C H…N and π–π interactions.

Proton NMR Studies of 5′-d-(TC) 3 (CT) 3 (AG) 3-3′—A Paperclip Triplex: The Structural Relevance of Turns

In this study, we present the results of structural analysis of an 18-mer DNA 5′-T 1C 2T 3C 4T 5C 6C 7T 8C 9T 10C 11T 12A 13G 14A 15G 16A 17G 18-3′ by proton nuclear magnetic resonance (NMR) spectroscopy and molecular modeling. The NMR data are consistent with characteristics for triple helical structures of DNA: downfield shifting of resonance signals, typical for the H3 + resonances of Hoogsteen-paired cytosines; pH dependence of these H3 + resonance; and observed nuclear Overhauser effects consistent with Hoogsteen and Watson-Crick basepairing. A three-dimensional model for the triplex is developed based on data obtained from two-dimensional NMR studies and molecular modeling. We find that this DNA forms an intramolecular “paperclip” pyrimidine-purine-pyrimidine triple helix. The central triads resemble typical Hoogsteen and Watson-Crick basepairing. The triads at each end region can be viewed as hairpin turns stabilized by a third base. One of these turns is comprised of a hairpin turn in the Watson-Crick basepairing portion of the 18-mer with the third base coming from the Hoogsteen pairing strand. The other turn is comprised of two bases from the continuous pyrimidine portion of the 18-mer, stabilized by a hydrogen-bond from a purine. This “triad” has well defined structure as indicated by the number of nuclear Overhauser effects and is shown to play a critical role in stabilizing triplex formation of the internal triads.

Inhibitors of NF-κB and AP-1 Gene Expression: SAR Studies on the Pyrimidine Portion of 2-Chloro-4-trifluoromethylpyrimidine-5-[N-(3‘,5‘-bis(trifluoromethyl)phenyl)carboxamide

We investigated the structure-activity relationship studies of N-[3, 5-bis(trifluoromethyl)phenyl][2-chloro-4-(trifluoromethyl)pyrimidin-5 -yl]carboxamide (1), an inhibitor of transcription mediated by both NF-kappaB and AP-1 transcription factors, with the goal of improving its potential oral bioavailability. Compounds were examined for cell-based activity, were fit to Lipinski's rule of 5, and were examined for potential gastrointestinal permeability using the intestinal epithelial cell line, Caco-2. Selected groups were substituted at the 2-, 4-, and 5-positions of the pyrimidine ring using solution-phase combinatorial methodology. The introduction of a fluorine in the place of 2-chlorine of 1 resulted in a compound with comparable activity. However, other substitutions at the 2-position resulted in a loss of activity. The trifluoromethyl group at the 4-position could be replaced with a methyl, ethyl, chlorine, or phenyl without a substantial loss of activity. The carboxamide group at the 5-position is critical for activity. If it was moved to the 6-position, the activity was lost. The 2-methyl analogue of 1 (81) showed comparable in vitro activity and improved Caco-2 permeability compared to 1.

Synthesis and anticancer evaluation of certain α-methylene-γ-(4-substituted phenyl)-γ-butyrolactone bearing thymine, uracil, and 5-bromouracil

Certain α-methylene-γ-(4-substituted phenyl)-γ-butyrolactone bearing thymine, uracil, and 5-bromouracil were synthesized and evaluated for their anticancer activity. These compounds demonstrated a strong growth inhibitory activity against leukemia cell lines. The anticancer potency for the substituents of the lactone C(γ)-phenyl is in an order of 4-Ph > 4-Cl, 4-Br > 4-Me, 4-NO 2 > 4-F. For the pyrimidine portion, 5-bromouracil is more potent than uracil and thymine.

Effect of selective cytosine methylation and hydration on the conformations of DNA triple helices containing a TTTT loop structure by FT-IR spectroscopy.

5-Methylcytosines have been introduced into triplex-forming-oligonucleotides and shown to extend the pH range over which a triplex forms with a homopurine-homopyrimidine tract of duplex DNA. As a host strand, an oligodeoxypyrimidine with a base sequence of 5'-d(TC)3T4(CT)3 ([CC]) was designed to form a hairpin triplex with a 5'-d-A(GA)2G ([AG6]) purine strand at acidic pH (Tsay, et al., (1995) J. Biomol. Str. Dyn., 13, 1235-1245). We here present results obtained by FT-IR spectroscopy concerning the conformation of the hairpin triplex as a function of the selective substitution of cytosines by 5-methylcytosines in the host strand. Namely, cytosines are substituted by 5-methylcytosines in either the 3'-pyrimidine portion ([CM]) or the 5'-pyrimidine portion ([MC]) or in both ([MM]) of the host strand. The acidic-induced transitions of the equimolar mixtures of the purine target with either of the four pyrimidine oligomers gives rise to different apparent pK values, i.e., [MM].[AG6] (6.2) > [MC].[AG6] (6.0) > [CM].[AG6] (5.7) > [CC].[AG6] (5.2) > single-stranded oligopyrimidines (4.6 +/- 0.2), indicating that cytosine methylation expands the pH range compatible with the hairpin triplex formation regardless of whether the substitution is in the 5'-pyrimidine (Hoogsteen) portion or in the 3'-pyrimidine (Watson-Crick) portion. Thermal denaturation profiles indicated that all the triplexes denatured in a monophasic manner in the pH range of 4.0 to 7.0, and that cytosine methylations in any position of the 16-base pyrimidine oligomer increase the stability of the hairpin triplex DNA. IR spectra recorded in D2O and H2O solutions revealed that cytosine methylation does not significantly influence the conformation of triplex DNA in solution, i.e., all the four triplexes accept a similar sugar conformation, and predominately take on a S-type sugar pucker with a relative proportion of two S-type sugars for one N-type. Furthermore, we also investigated the effect of relative humidity (RH) on the conformation of triplex MC.AG6 in hydrated films, and found that the conformational change induced by the decrease of RH, from predominant S-type to primary N-type sugar pucker, might first occur in the purine strand at 86% RH.

Lazaroids improve the survival of grafted rat embryonic dopamine neurons.

In rodent models of Parkinson disease in which transplants of dissociated rodent and human embryonic mesencephalic tissue, rich in dopamine neurons, have been studied, only 5-20% of the dopamine neurons survive the implantation procedure. We have investigated the effects of inhibiting free radical generation with two lazaroids, U-74389G and U-83836E, on the survival of embryonic rat dopamine neurons. U-74389G is a 21-aminosteroid, and U-83836E combines the piperazinyl pyrimidine portion of 21-aminosteroids with the antioxidant ring of alpha-tocopherol. In an initial study, we found that the lazaroids markedly prolonged the period after tissue dissociation that an embryonic mesencephalic cell suspension exhibits high cell viability in vitro, as assessed by using a dye exclusion method. In a second series of experiments, addition of lazaroids to dissociated mesencephalic graft tissue increased the yield of surviving rat dopamine neurons 2.6-fold after implantation in the dopamine-denervated rat striatum. The improved survival correlated with an earlier onset of graft-induced functional effects in the amphetamine-induced rotation test. Thus, inhibition of free radical generation can significantly increase the yield of grafted embryonic dopamine neurons. Addition of lazaroids to the graft preparation is a relatively simple modification of the transplantation protocol and could readily be applied in a clinical setting. Moreover, since iron-dependent lipid peroxidation has been suggested to play a role in the death of nigral dopamine neurons in Parkinson disease and lazaroids are particularly potent inhibitors of such processes, the findings may have implications for the pathogenesis of this disease.

The structure of flavocytochrome c sulfide dehydrogenase from a purple phototrophic bacterium.

The structure of the heterodimeric flavocytochrome c sulfide dehydrogenase from Chromatium vinosum was determined at a resolution of 2.53 angstroms. It contains a glutathione reductase-like flavin-binding subunit and a diheme cytochrome subunit. The diheme cytochrome folds as two domains, each resembling mitochondrial cytochrome c, and has an unusual interpropionic acid linkage joining the two heme groups in the interior of the subunit. The active site of the flavoprotein subunit contains a catalytically important disulfide bridge located above the pyrimidine portion of the flavin ring. A tryptophan, threonine, or tyrosine side chain may provide a partial conduit for electron transfer to one of the heme groups located 10 angstroms from the flavin.

Anthranilic acid release in adenosine-inhibited cultures of Saccharomyces cerevisiae and its inhibition by thiamin

Adenosine, at 1 mM concentrations or above, was found to have a fungistatic effect on Saccharomyces cerevisiae. A substance with amethyst fluorescence wa was detected in the medium of adenosine-inhibited cultures of S. cerevisiae. This compound was isolated and physicochemically identified as anthranilic acid. Both the inhibition of growth and release of anthranilic acid induced by adenosine were abrogated by thiamin or by the pyrimidine portion of thiamin, 2-methyl-4-amino-5-hydroxymethyl-pyrimidine (hydroxymethyl-pyrimidine); the latter was found to restore intracellular thiamin content that had been reduced by adenosine. It was demonstrated that effects of thiamin and hydroxymethylpyrimidine on S. cerevisiae cultured with adenosine resulted from their inhibition of adenosine uptake by growing yeast cells.

Anthranilic acid release in adenosine-inhibited cultures ofSaccharomyces cerevisiaeand its inhibition by thiamin

Adenosine, at 1 mM concentrations or above, was found to have a fungistatic effect on Saccharomyces cerevisiae. A substance with amethyst fluorescence wa was detected in the medium of adenosine-inhibited cultures of S. cerevisiae. This compound was isolated and physicochemically identified as anthranilic acid. Both the inhibition of growth and release of anthranilic acid induced by adenosine were abrogated by thiamin or by the pyrimidine portion of thiamin, 2-methyl-4-amino-5-hydroxymethyl-pyrimidine (hydroxymethyl-pyrimidine); the latter was found to restore intracellular thiamin content that had been reduced by adenosine. It was demonstrated that effects of thiamin and hydroxymethylpyrimidine on S. cerevisiae cultured with adenosine resulted from their inhibition of adenosine uptake by growing yeast cells.

ChemInform Abstract: Nonenzymatic Synthesis and Properties of 5‐Aminoimidazole Ribonucleotide (AIR). Synthesis of Specifically 15N‐Labeled 5‐Aminoimidazole Ribonucleoside (AIRs) Derivatives.

AbstractThe chemical synthesis of 5‐aminoimidazole ribonucleotide (AIR) (III) from the ribonucleoside (AIRs) (I) is described; the former is the precursor of the purine ribonucleotides in vivo in both prokaryotic and eukaryotic systems as well as the biosynthetic precursor of the pyrimidine portion of vitamin B1 in certain prokaryotic organisms.

Nonenzymatic synthesis of 5-aminoimidazole ribonucleoside and recognition of its facile rearrangement.

5-Amino-1-beta-D-ribofuranosylimidazole 5'-monophosphate (AIR, 1) is the ubiquitous precursor to the purine ribonucleotides in vivo, and it serves as the biochemical precursor to the pyrimidine portion of thiamin (vitamin B1) in certain prokaryotic organisms. The corresponding ribonucleoside (AIRs, 5b) was prepared via chemical (nonenzymatic) synthesis from 5-amino-1-beta-D-ribofuranosylimidazole-4-carboxamide. The tri-O-acetylated derivative of AIRs (5a) was also prepared, and it was shown to undergo a facile ring transformation in aqueous pH 7 buffer to afford N-(imidazol-4-yl)-2,3,5-tri-O-acetyl-D-ribofuranosylamine as a 1:2 mixture of alpha and beta anomers (6a). Under similar conditions, compound 5b affords the corresponding unprotected beta-ribonucleosides 6b. This Dimroth-type ring transformation reaction of 5 to 6, which occurs primarily in neutral aqueous solution, may be responsible for the previously reported lability of AIRs and its derivatives. It may also have relevance to the postulated early biotic pathway to the 9- and 3-substituted purine nucleotide components of an all-purine biopolymer.

Aldehyde oxidase from rabbit liver: Specificity toward purines and their analogs

Aldehyde oxidase (EC 1.2.3.1) plays a role in the oxidation of aromatic heterocyclic compounds ingested by some higher vertebrates. To better understand this function, the specificity of the rabbit liver enzyme toward purines and their analogs was quantitatively studied. The chemical nature of the 6-substituent of purine markedly influenced substrate efficiency ( V max K m ). Substituents that were hydrophobic were generally favorable. There was a correlation between the degree of hydrophobicity and the tightness of binding. 6-Substituents that were strongly electron-withdrawing also enhanced substrate efficiency. 6-Hydroxy and 6-amino substituents virtually obliterated substrate activity. In contrast, 2-hydroxypurine and 2-aminopurine were efficiently oxidized. 2,6-Disubstitution of purine was much less favorable than either 2- or 6-monosubstitution. N-Substitution of purines enhanced substrate efficiency in many cases. The typical order of preference for 9-substituents was 2′-deoxyribofuranosyl > ribofuranosyl > arabinofuranosyl > H. Acyclic nucleosides (9-[(hydroxy-alkyloxy)methyl]purines) were usually more efficient substrates than were 2′-deoxyribonucleosides. The kinetic constants of a variety of purine analogs revealed that the pyrimidine portion of the purine ring system was a more important determinant of substrate activity than the imidazole portion. The efficient oxidation of a variety of nucleosides suggests that detoxification of naturally occurring nucleoside analogs might be an important aspect of the physiological role of this enzyme. Overall, the data presented serve as a guide for predicting the susceptibility of heterocycles to oxidation by this enzyme.

Cyclic GMP directly regulates a cation conductance in membranes of bovine rods by a cooperative mechanism.

Cyclic GMP causes the release of endogenous Ca2+ from rod outer segments, whose plasma membrane has been made permeable, or from isolated discs. Approximately 11,000 Ca2+ ions are released per disc at saturating concentrations of cyclic GMP. The velocity and the amplitude of the release of Ca2+ are dependent on the concentration of cyclic GMP. The maximal rate of the Ca2+ efflux is approximately 7 X 10(4) Ca2+ ions s-1 rod-1. The Ca2+ release by cyclic GMP is independent of light. The activation of the efflux occurred within a narrow range of the cyclic GMP concentration (30-80 microM) and does not obey a simple Michaelis-Menten scheme. Instead, the kinetic analysis of the Ca2+ efflux suggests that a minimum number of 2 molecules of cyclic GMP activates the ion conductance in a cooperative fashion. The release of Ca2+ by cyclic GMP requires a gradient of Ca2+ ions across the disc membrane. If the endogenous Ca2+ gradient is dissipated by means of the ionophore A23187, the release of Ca2+ by cyclic GMP is abolished. Ca2+ is released by analogues of cyclic GMP which are either modified at the 8-carbon position of the imidazole ring or by the deaza-analogue of cyclic GMP. Congeners of cyclic GMP which are modified at the ribose, phosphodiester, or pyrimidine portion of the molecule are ineffective. The hydrolysis of cyclic GMP by the light-regulated phosphodiesterase of rod outer segments is not a necessary condition for the Ca2+ release because 8-bromo-cyclic GMP, a congener resistant to hydrolysis, is a more powerful activator of the release than cyclic GMP itself. Ca2+ release by cyclic GMP is inhibited by organic and inorganic blockers of Ca2+ channels. The l-stereoisomer of cis-diltiazem blocks the release of Ca2+ at micromolar concentrations, whereas the d-form is much less effective. These results suggest that disc membranes contain a cationic conductance which is permeable to Ca2+ ions and which is regulated through the cooperative binding of at least 2 molecules of cyclic GMP to regulatory sites of the transport protein. By this mechanism, subtle changes in the concentration of cyclic GMP could promote large changes in the flux of Ca2+ ions across the disc membrane.

Examination of the structure of an unknown green fluorescent compound, compound G2, accumulated in non-growing cells of Eremothecium ashbyii by the addition of dimeric diacetyl.

The addition of dimeric diacetyl to Eremothecium ashbyii caused simultaneous accumulation of two green fluorescent compounds with the inhibition of riboflavin formation in non-growing cells. One compound, referred to as Compound G1, was identified as 6, 7-dimethyl-8-ribityllumazine as reported previously and the other is referred to as Compound G2. The latter compound was considered to be 6-methyl-7-(2-hydroxy-2-methyl-3-oxobutyl)-8-ribityllumazine because a violet fluorescent compound, 6-methyl-7-hydroxy-8-ribityllumazine was derived from Compound G2 in the presence of p-quinone and because the known action mechanism of dimeric diacetyl as a trapping agent of possible intermediates in the biosynthetic pathway. The results indicate that an immediate intermediate to 6, 7-dimethyl-8-ribityllumazine in the biosynthetic pathway of riboflavin is 4-ribitylamino-5-amino-2, 6-dihydroxypyrimidine, whose pyrimidine portion is derived by the elimination of a dimeric diacetyl fragment from the molecule of the isolated lumazine.

Pyrimidine metabolism in seeds and seedlings of jack pine (Pinus banksiana)

Jack pine seedlings rapidly converted radioactive thymine-6-3H to dihydrothymine, β-ureidoisobutyric acid, and β-aminoisobutyric acid. Autoradiographs of hypocotyl cells showed radioactivity largely in the cytoplasm. A small percentage of radioactivity from thymine-2-14C was salvaged via uracil for the synthesis of RNA. DNA was not labelled significantly. The occurrence of a mechanism to utilize thymidine-methyl-3H for DNA synthesis was detected at low levels in germinating seedlings but was absent in seeds up to 16 h imbibition. Among pyrimidines tested thymidine was the most specific for the labelling of DNA and radioactivity was detected almost entirely over nuclei.The occurrence of an active pathway for the de novo synthesis of the pyrimidine portions of nucleic acids was demonstrated by the use of orotic acid-4-14C. Native DNA and all major types of RNA were labelled and radioactivity resided in pyrimidines. Synthesis of DNA involved the conversion of pyrimidine ribonucleotides to deoxyribonucleotides.

Uridine diphosphate glucose dehydrogenase of calf liver: Properties and inhibition characteristics with uridine diphosphate xylose analogues

UDP-glucose dehydrogenase of calf liver dissociated in guanidine-HCl into six subunits. The number of reactive sulfhydrylgroups of native and guanidine-HCl-treated enzyme was found to be 20 ± 1 and 46 ± 1, respectively, per mole of native enzyme. A crude preparation of UDP-glucose pyrophosphorylase from yeast was used for the preparation of 5-hydroxyuridine diphosphate xylose and 5-aminouridine diphosphate xylose. 5,6-Dihydrouridine diphosphate xylose was prepared by catalytic reduction of UDP-xylose. The nature of the inhibition produced by UDP-xylose or its analogues was similar with respect to either UDP-glucose or NAD +, the two substrates for the enzyme. Hydrogenation of the 5,6-double bond or substitution of a hydroxyl group at the C-5 position of the pyrimidine portion of UDP-xylose decreased its inhibitory activity. Substitution of an amino group at the C-5 position, however, did not alter the activity of the allosteric inhibitor.

X-Ray Analysis of Thiamine Propyl Disulfide

The crystal structure of thiamine propyl disulfide (TPD) was determined by the three dimensional Patterson superposition method and by application of least squares treatment and Fourier syntheses. The unit cell is monoclinic, space group P21/a, with a=21.04, b=11.35, c=8.53 A and β=101°36'. The propyl disulfido group has been found to lie cis to the N-formyl group, the result being consistent with the hitherto proposed structure for TPD. The structure of the pyrimidine portion was discussed in comparison with the results of earlier workers. The stereochemical structure of TPD and a unique hydrogen bonding in the crystal were also discussed.

Base-Catalyzed Hydrolysis of Flavins: Effect of Amines and the Particular Role of Position 3

The base-catalyzed hydrolysis of flavins was investigated in 50 percent water-methanol with excess sodium hydroxide or excess amines at different temperatures. Rate constants for the pseudo first-order cleavage of the pyrimidine portion of the isoalloxazine systems were measured and with sodium hydroxide shown to increase from lumiflavin to 3-ethyllumiflavin to 3-carboxyamido(phenylalanyl)lumiflavin. With amines, the size as well as basicity is of prime importance since the general order in efficacy of the nucleophile for hydrolysis of flavins is ethylamine > diethylamine > triethylamine > benzylamine > tributylamine. In general, alkylation of the 3-imino function of the flavin increases its lability because of inability to ionize and undergo resonance stabilization of the anionic tautomers. Moreover, the special case where a 3-carboxyamido function is present allows intramolecular polarization, <i>via</i> hydrogen bonding of the amide hydrogen to 4-carbonyl oxygen, which causes a marked increase in hydrolysis rate in sodium hydroxide. From the temperature dependencies of the rates of hydrolysis, the calculated values for energies of activation indicate the decrease expected upon 3-alkylation and further upon intramolecular enhancement with the 3-carboxyamido group. The calculated values for entropies of activation are large and negative in accord with considerable loss of degrees of freedom in the activated complexes formed in the bimolecular mechanism.

Metabolic disorders in thiamineless dwarf strains of Staphylococcus aureus.

SUMMARY: Dwarf colony variants of Staphylococcus aureus have been described as a causative agent of bovine mastitis in Israel. These strains were reported as auxotrophic for thiamine, being unable to synthesize the vitamin from its pyrimidine and thiazole moieties. In the present paper it is shown that strains with two different deficiencies are involved. One type is unable to concentrate the thiazole moiety and the other is unable to phosphorylate the pyrimidine portion of the vitamin. The results indicate the existence of independent permeases in S. aureus for these thiamine moieties.