1 School of Biological Sciences, University of Wales Bangor, Gwynedd LL57 2UW, UK; 2 Department of Pharmaceutical Technology and Biochemistry, Technical University of Gdaásk, Gdaásk, 80-952, Poland
Received 3 December 2002; returned 18 January 2003; revised 21 January 2003; accepted 21 January 2003
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Abstract |
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Keywords: cell wall biosynthesis, peptide transport, molecular modelling, molecular recognition
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Introduction |
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FMDP-peptides have been shown to inhibit various bacterial species, including Escherichia coli (ATCC 9637), Shigella sonnei 433, Staphylococcus aureus 209P, Bacillus pumilus 1697, Bacillus subtilis3,6,8,9 and also fungi, e.g. Candida albicans.6,9 However, the inhibition assays allowed only a rough ranking of relative antibacterial activities, e.g. for E. coli MICs of 100 or >200 mg/L,3 or the inhibitory effects of a single amount of each inhibitor using disc diffusion assays.8 The rate-limiting step for antimicrobial activity is generally uptake, although for B. subtilis uptake rates were generally faster than the intracellular hydrolysis rates that liberated free FMDP.8 Furthermore, certain of these compounds have demonstrated chemotherapeutic activity in a mouse model of generalized candidiasis,10,11 showing them to be potentially valuable, broad-spectrum antimicrobial compounds.
Molecular modelling of small peptides (two to five residues) derived from protein hydrolysis has allowed identification of the conformational forms they adopt in solution, and revealed how various peptide transporters have evolved complementary specificities to recognize different conformational types as ligands.12,13 From evaluation of the structural and electronic features needed for this ligand recognition, individual molecular recognition templates (MRTs) for the microbial peptide transporters have been defined.1417 The following features are involved in defining MRTs: (i) charged N-terminal -amino and C-terminal
-carboxylate groups, permitting charge and hydrogen-bond donor and acceptor properties; (ii) backbone torsion angles psi, phi and omega; (iii) chirality at
-carbons; (iv) NC distance between terminal amino and carboxylate groups; (v) chi space torsions and size of side chains; (vi) H-bond donor and acceptor properties of peptide bond atoms; and (vii) charge fields around N-terminal
-amino group and C-terminal
-carboxylate. To define an MRT, each of these seven features must be of a certain type, e.g. positively-charged N-terminal
-amino group or L-chirality, or value, e.g. psi torsion angle of 150°. Knowing these MRTs, compounds can be modelled to assess their match to these parameters and thus their potential as putative transport ligands.
In the present study, structureactivity relationships have been explored for FMDP peptides using E. coli. The substrate specificities of the generic peptide transporters Opp, Dpp and Tpp have well-characterized features that allow them to be distinguished from each other, but they also share certain substrate recognition features, so that some peptides can be taken up by more than one transporter.14,17 Dpp and Opp are typical ABC transporters energized by ATP; each comprises four membrane proteins plus a periplasmic peptide binding protein DppA or OppA, respectively.17 In contrast, Tpp lacks a binding protein and comprises a single membrane protein that is energized by a proton-motive force.17 To be transported by Dpp or Opp, ligands must be recognized and bound by their respective binding proteins. A strong correlation exists between the affinity of ligands for these binding proteins and uptake rates through the respective transporters.12,14,16,18 Thus, compounds can be evaluated as potential transport ligands by measuring their abilities to compete for binding of radioactively labelled ligands to DppA and OppA, which is not only more convenient than directly assaying transport but in requiring less sample is particularly convenient when little material is available, as with many of the peptidomimetics here. For antibacterial compounds, such binding studies can complement inhibition assays. Here, structureactivity results have been obtained using three approaches: first, antibacterial activities against isogenic strains of E. coli that possess different complements of the peptide transporters; secondly, peptide binding to purified peptide binding proteins; and thirdly, molecular modelling to derive structural information on the conformations adopted by the peptidomimetics in solution. The results should aid the design of antimicrobial peptidomimetic prodrugs of glutamine analogues.
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Materials and methods |
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Peptidomimetics containing FMDP, N3-(fumaramoyl)-L-2,3-diaminopropanoic acid (FCDP) or N4-(4-methoxyfumaramoyl)-L-2,4-diaminobutanoic acid (FMDB) were synthesized as described previously.3,8 The iodinated peptide ligands, Gly[125I]Tyr and [125I2]TyrGlyGly were prepared as described previously.18 The synthetic inhibitor, alafosfalin [Ala-L-1-aminoethylphosphonic acid; AlaAla(P)] was a gift from Roche Products, Welwyn Garden City, UK.14,18
Determination of the antibacterial activities of peptidomimetics
Peptidomimetics were tested using the following strains, the isolation and characterization of which were described previously:18 a parental strain, E. coli K-12 Morse 2034 (trp, leu) (CGSC 5071), which is a wild-type with respect to Opp, Dpp and Tpp; and three isogenic mutants: (i) strain PA0183 (opp), derived from strain Morse 2034, in which the deletion extends from tonB to tdk; (ii) strain PA0333 (
opp,dpp) derived from PA0183; and (iii) strain PA0410 (
opp,tpp) also derived from PA0183. Inhibition was determined using the following disc diffusion assay, adapted from recommended procedures.19 Bacteria were grown in minimal A medium18 supplemented with 0.5 mM L-Leu, 0.2 mM L-Trp, 0.05 mM FeCl3 and 0.4 % (w/v) D-glucose at 37°C with rotary shaking. Soft agar overlays (0.6 % w/v agar), similarly supplemented with Leu, Trp and glucose, were inoculated with exponential-phase bacteria to give
1 x 107 cells/mL, to allow semi-confluent growth, and used to overlay a plate (1.5 % w/v agar) of the same composition. Typically, six filter paper discs were placed on the surface, 10 µL amounts of different concentrations of the peptidomimetics were added, giving a range of 30300 nmol, and the plates were incubated at 37°C. After 17 h incubation, the appearance of any inhibition zones was noted and their diameters measured. Antibacterial activity was expressed as the amount of inhibitor (nmol) required to produce an inhibition zone of 25 mm and was determined from semi-logarithmic plots of log10 concentrations of inhibitor versus inhibition zone diameters. These isogenic strains all have the same rates of growth in the above liquid medium and on these agar plates, so zone sizes are not susceptible to different growth rates18 (data not shown).
Assays for peptide binding to DppA and OppA
DppA and OppA were isolated from strain Morse 2034 by an osmotic shock procedure, purified to homogeneity by sequential use of ion-exchange chromatography, freed from any endogenous ligands using reverse-phase chromatography, lyophilized and stored at 20°C, as described.14,18 Competitive filter-binding assays were carried out using Gly[125I]Tyr as the radioactive ligand for DppA, and [125I2]TyrGlyGly for OppA, as described previously.18 All competitor peptidomimetics were tested at a molar ratio of 10:1 relative to the ligands, and some were tested at other molar concentrations. As control competitors, AlaAla was used with DppA and AlaAlaAla with OppA. Controls were performed in which either the binding protein or a competing peptide was omitted.
Molecular modelling of peptides and peptidomimetics
Conformational analysis of zwitterionic peptides and peptidomimetics was carried out using SYBYL software (Tripos Inc., St Louis, MO, USA), in a similar manner to that described in detail previously.1216,20,21 Briefly, starting structures were assigned appropriate atom types and charges, and were energy-minimized before being subjected to random searches (25k search cycles) with an absolute energy cut-off of 40 kcal/mol, using a distance-based dielectric constant of 80 to simulate a water environment. Unique conformers were distinguished by setting a root mean square threshold of 0.2 Å, together with chirality checking. The computed collection of conformers for each compound was first ordered according to energy, before the percentage contribution of each conformer was determined using a Boltzmann distribution. For each dipeptide conformer, backbone torsion angles, psi () (Tor2), omega (
) (Tor3) and phi (
) (Tor4) were measured, together with relevant side-chain torsions, chi (
) and the distance (NC) between the nitrogen of the N-terminal
-amino group and
-carbon of the C-terminal
-carboxylate group. For tripeptide conformers, analogous measurements were made, together with those for (
i) Tor6, (
2) Tor7 and (
i+1) Tor8 (Figure 1). For each dipeptide compound,
and
torsional space for the peptide unit was divided up into 36 10° sectors and the percentage contributions of conformers having particular
combinations were aggregated. Molecular modelling of peptides has previously indicated that conformers adopt a limited set of torsional combinations, which have been classified by dividing
and
conformational space into 12 30° sectors designated A1 to A12 (0° to ±180°) and B1 to B12 (0° to ±180°), respectively.12,1416 In this way, particular conformational forms can be defined by reference to their location within the conformational grid, e.g. A7B9. Analogous procedures were adopted for tripeptides.16 A complete profile of the conformers of any compound can be displayed using 3D pseudo-Ramachandran plots (3DPR) that relate cumulative percentage of conformer to backbone
and
torsions.12,1416,20
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Results |
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The antibacterial activities of the peptidomimetics were tested against the parent strain M2034 and peptide-transport mutants of E. coli (Table 1). Because only limited amounts of the synthetic peptidomimetics were available, all compounds were initially tested against the parent strain and those that did not give a measurable level of inhibition, e.g. Gly-FMDP (Table 1), were not tested with the mutants. In addition, tests with the mutants were performed on the basis of their substrate specificities, e.g. Opp favours peptides with three to five residues and transports dipeptides poorly, whereas Dpp and Tpp transport di- and tripeptides only but with different specificities.17,18,22 Alafosfalin, a well-characterized, synthetic, antimicrobial peptide prodrug (smugglin),7,22 was used (0.5 nmol) as a control to check reproducibility of assay procedures: for example, an inhibition zone of 18.6 ± 0.9 mm was found for all assays with strain M2034.
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Active Xaa-FMDP analogues can generally be transported by both Dpp and Tpp, i.e. strains PA0410 and PA0333 were inhibited, with activity against the former usually being greater (Table 1). Their activities against strains M2034 and PA0183 indicated that uptake by Opp also contributed to their overall antibacterial activity (Table 1). For the three oligopeptides, inhibition of wild-type M2034 and no antibacterial activity against PA0183 (Opp,Dpp+,Tpp+) indicate that their uptake occurs exclusively by Opp (Table 1). Thus, although both Dpp and Tpp can transport folded conformers of many natural tripeptides, these particular tripeptide mimetics are effectively recognized only by Opp, which recognizes elongated conformers.14,17,22
Binding of peptidomimetics to DppA and OppA
Several trends can be observed from the relative abilities of the dipeptide analogues to compete for binding with Gly[125I]Tyr to DppA (Table 2). Xaa-FMDP dipeptides are generally better competitors than FMDP-Xaa dipeptides; this was exemplified specifically when comparing Leu-FMDP and Met-FMDP with FMDP-Leu and FMDP-Met. Most Xaa-FMDP dipeptides showed broadly similar levels of competitive ability, although no activity was detectable for Gly-FMDP and FMDP-FMDP. These results are all in broad agreement with the relative inhibitory activities of these compounds (Table 1), indicating that transport rates appear to be the main determinant of antibacterial activities. AcNva-FMDP also showed no competitive activity (Table 2), in accordance with the requirement for a protonated N-terminal -amino group for ligand binding to DppA and transport by Dpp.17,18,22 Consequently, lack of binding and presumed transport is sufficient explanation for its failure to inhibit (Table 1). In addition to the results for a ligand:competitor ratio of 1:10 (Table 2), Nva-FMDP and Nva-FMDB were also assayed at 1:1 and 1:5 ratios and found to inhibit 73% and 93%, and 89% and 100%, respectively. Similarly, FCDP-Ala and FCDP-Met are better competitors than FMDP-Ala and FMDP-Met (Table 2), compatible with them being well transported and implying that the very poor antibacterial activity of the FCDP-containing dipeptides (Table 1) relates to the lower enzyme inhibition of the FCDP warhead compared with FMDP.1
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Molecular modelling of peptides and peptidomimetics
Molecular modelling of various classes of natural and synthetic peptides has allowed identification of the bioactive conformational forms recognized by different peptide transporters and peptidases, and provided an understanding of structureactivity relationships for substrates of these proteins.1214,16,20 Backbone torsions are a very important feature of MRTs for peptide transporters.12,1417 For the peptide bond only the trans form is recognizable.24,25 A variety of values can be adopted by
and
torsions. Dipeptides mostly adopt Tor2 of A4 (50° to 85°), A7 (+140° to 175°) and A10 (+50° to +85°), and Tor4 of B2 (+40° to +85°), B9 (50° to 95°) and B12 (130° to +175°) (Figure 1), and these torsions help to determine the ligand specificities of peptide transporters.12,1417 Thus, in the present case, conformational analysis of the peptidomimetics can provide an estimate of the extent to which the conformer repertoire of each matches the MRTs for substrate(s) of each peptide transporter. This evaluation of the peptidomimetics as putative ligands of Dpp, Tpp and Opp can be related to the ligand binding data, and, in turn, to the antibacterial results, so as to explore the basis for structureinhibition relationships. Random search conformational analysis for each peptidomimetic and related Xaa-Gln dipeptides showed they occurred as several hundred distinct conformers (results not shown), comparable to the finding for most other natural peptides.15,16 The various conformers for any one compound, e.g. 768 for Val-FMDP, are distinguishable either by having different combinations of backbone torsions (
,
and
), or having comparable backbone torsions but different orientations of side chains (
torsions). Using a random search procedure, the minimum energy conformation identified may or may not be the actual global minimum conformer and it may or may not be a bioactive form. However, this is not of great importance here, for what is critically important when considering molecular recognition of flexible molecules is to consider the whole population of conformational forms and to be able to estimate with good precision the percentage of conformers that match MRTs.13,21 Thus, there is a direct correlation between the proportion of conformers that match a particular MRT and the bioactivity of the compound, e.g. affinity to the ligand, transport rate, etc.1214,20,24
For these compounds, the minimum energy form typically accounts for 830% of the total conformer pool, and there are broad similarities between the energies of Xaa-Gln peptides and their respective peptidomimetics. Because all the peptidomimetics comprise L-amino acids with charged N- and C-terminal groups, when the minimum energy conformers have backbone torsions that match those for peptide transporter MRTs they can be considered as putative ligands. For each peptidomimetic, data for the percentage of its conformers having backbone torsions and lengths matching the MRTs for Dpp, Tpp and Opp are shown in Table 3. Three-dimensional pseudo-Ramachandran plots showing the complete conformer distribution for representative inhibitors are shown in Figure 2. From these results, several general trends can be seen regarding structureactivity relationships for the inhibitors. For the Xaa-FMDP compounds, there is broad similarity in their conformer profiles with those of the related Xaa-Gln dipeptides, and they have comparable proportions of conformers matching the different MRTs, e.g. Tyr-FMDP (65%, 18%) and Tyr-Gln (64%, 17%); Val-FMDP (55%, 19%) and Val-Gln (54%, 23%); Leu-FMDP (15%, 55%) and Leu-Gln (21%, 50%) for Dpp and Tpp, respectively. These percentage values for the Xaa-FMDP peptidomimetics are generally in broad agreement with their relative antibacterial activities (Table 1). The antibacterial activity of these compounds appears mainly to arise from uptake by Dpp, in accordance with them having a higher proportion of conformers that match the MRT for Dpp and, also, as a result of the greater inherent transport capacity of Dpp.17,22 For AcNva-FMDP, its inactivity can be attributed to substitution of its N-terminal
-amino group, which must possess a positive charge to achieve ligand binding to Dpp, Tpp and Opp. For both DppA and OppA, this involves interaction of the positive charge on the
-amino group with the side-chain carboxylate of an aspartic acid residue.12,17,22 The antibacterial activity of Gly-FMDP is extremely poor, which is in accordance with the general finding that any dipeptide containing Gly shows low transport activity by Dpp, Tpp and Opp.17 This arises from several causes. First, a high proportion of their conformers have cis
bonds, which are not recognized.14,24,25 Secondly, the small side chain allows greater flexibility of their
and
torsions so they can adopt a wider array of conformational forms, very few of which match well the backbone torsions favoured for recognition by the transporters. The need to define weightings for the relative contributions of such conformers with torsions outside the optimum range (Table 3) still needs to be adequately addressed.15,24 Significantly, the most effective inhibitor of mutant PA0333 (Tpp+) is Leu-FMDP (Table 1) and this accords well with the finding that (but for Lys-FMDP) it has the highest proportion of conformers (55%) that match precisely the backbone torsions of the MRT for Tpp (Table 3, Figure 2). Thus, Lys-FMDP alone appears anomalous in the relationship between its antibacterial activity and conformer distribution based on backbone torsions, and an explanation for this is considered below.
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Results with Lys-FMDP provide a further test of the present approach of using conformational analysis to probe structureactivity relationships. Lys-FMDP inhibited the parent strain and mutants PA0183 (Dpp+,Tpp+) and PA0410 (Dpp+) but showed no activity against mutant PA0333 (Tpp+) (Table 1), indicating that its uptake is predominantly by Dpp, a little by Opp, but not at all by Tpp. These results appear at odds with the finding that the majority of its conformers have backbone torsions that match the MRT for Tpp (Table 3, Figure 2). However, particular properties of a side chain can also markedly influence the molecular recognition of peptides, as illustrated above regarding size, and described earlier for certain side chains that are charged and have particular torsions.1214 Examination of the molecular structures of A4B9, A4B12 conformational forms of Lys-FMDP, which have the lowest energies and are most abundant, indicates that both side chains are contiguous and fully extended in which trans
torsions predominate, allowing stabilizing interactions that are unique to these conformational forms (Figure 3). Thus, Lys-FMDP is quite atypical, and in its A4(B9,B12) conformers the side chains appear as a fused unit. In contrast, natural dipeptides having charged or polar N-terminal residues, e.g. LysGln has mainly A7 conformers in which the side chains are separated.13,15
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Discussion |
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Examples of the glutamine analogue peptidomimetics studied here have been shown previously to have broad-spectrum antimicrobial activities and chemotherapeutic activity in a mouse model of candidiasis. In addition, extensive data have been obtained previously on their transport and cleavage by microorganisms, but these have not provided an understanding of the structural basis for their individual activities.811 In the present study, the relative antibacterial activities of a series of these peptidomimetics against isogenic strains of E. coli differing only in their complement of peptide transporters, has shown the extent to which the different transporters recognize and absorb the various compounds. Each strain has an identical complement of peptidases, so intracellular cleavage of peptidomimetics is comparable in each. Given the limited availability of compounds, the disc diffusion assay was chosen as a suitable test to obtain quantitative antibacterial data that was dependent upon the uptake of the various compounds through distinguishable peptide transporters. In this assay, a query can always be raised regarding possible variations in diffusion rates of compounds and whether hypothetically this may influence zone sizes, but only in very rare cases would it be possible to measure diffusion with any precision in these assays so as to check this speculation. In general, diffusion may be influenced by mass, shape, pKa, hydrophobicity, etc., but for compounds as closely related to each other as these it seems improbable that any such variations could have any significant effect on diffusion rates. Certainly, no correlation exists between the sizes of the compounds and their zone sizes. Thus, as is usual with this assay, it appears sensible to accept the inhibition zones as directly reflecting antimicrobial activities. This approach is supported by the correlations between the antibacterial assays and the results obtained from measurements of the relative affinities of the compounds for the peptide-binding proteins DppA and OppA, which are the components responsible for ligand specificities in the Dpp and Opp transporters. In general, there was good agreement between ligand binding and antibacterial activity; and this same conclusion has been reached previously for other antimicrobial peptidomimetics.17,18
Explanation of the structural basis for these activities was provided by comparison with results of molecular modelling of the various compounds. These peptidomimetics are the first example of a series of antimicrobial compounds for which such an analysis has been attempted. They are flexible molecules that exist as a variety of conformers in solution, and to understand how this determines their molecular recognition, transport, antibacterial activities, etc. it is necessary to analyse the complete repertoire of conformers for each compound. Previous analysis of natural peptide substrates has identified the important structural and electronic features, e.g. charged termini, particular backbone torsions, L-chirality, etc., which must be possessed by a compound for it to be well-recognized by peptide transporters in microorganisms and other species. Thus, Dpp recognizes dipeptide conformers with torsions of A7 paired with B9 and B12, whereas Tpp recognizes a different set of dipeptide conformers with torsions of A4 and A10 paired with B9 and B12; Dpp and Tpp also recognize folded tripeptide conformers with the above torsional values and appropriate NC distances. Opp recognizes elongated conformers of tri- and higher oligopeptides (and to a lesser extent dipeptides) with A7 combined with B9 and B12 and NC distances greater than 5.5 Å.12,1416 These conclusions have been corroborated by studies of bound peptide ligands in crystal structures of DppA and OppA.16,3033 These structural parameters have been incorporated into definitions of MRTs for each microbial transporter, which also appear to relate to mammalian and plant transporters.12,13,16,24 Thus, in principle, comparison of the extent to which the conformers of the peptidomimetics match the MRTs allows estimation of their transport rates and antibacterial activities.
For most of the Xaa-FMDP compounds and the tripeptidomimetics, structureactivity relationships were clear, with preferences for Dpp, Tpp or Opp, and thus antibacterial activities against the different strains, being related to the proportions of conformers that adopted the particular backbone torsions recognized by each transporter. However, a particular strength of this study is how it has provided insight into the influence of side chain properties, e.g. overall size and torsions, on molecular recognition. Thus, the lower bioactivities of FMDP-Xaa compounds compared with the corresponding Xaa-FMDP analogues, which are not explicable in terms of their backbone torsions, can be related to the dimensions of the side chain of FMDP. These values can be related to results of X-ray crystal studies on DppA and OppA, which show that they have water-filled pockets to accommodate the variety of side chains found in natural peptides. These have evolved so that different numbers of water molecules are displaced depending on the size of the side chain.3033 DppA possesses two pockets: thus, for dipeptide ligands, the N- and C-residue side chains are readily accommodated.31 For DppA (and Tpp) to bind tripeptides, a tripeptide conformer must be folded so that its
i1 and
i torsions and the locations of its N- and C-terminal charged groups mimic those in dipeptide ligand conformers. This requires that the C-terminal region (second peptide bond and the central and C-terminal side chains) is recognized as a unit that fits into the C-terminal pocket.12,1416 To achieve this, the C-terminal pocket needs to be larger than the N-terminal pocket and this is exactly what has been found.31 Consequently, the greater size of FMDP compared with the largest protein amino acids makes it likely that it is not easily accommodated in the N-terminal water-filled pocket but it can be in the C-terminal one. This observation provides a plausible explanation for the much poorer affinities of the FMDP-Xaa compounds compared with the Xaa-FMDP analogues for DppA (Table 2) (and implicitly for Tpp) and for their lower antibacterial activities (Table 1). FMDP-FMDP showed no binding to DppA and lacked measurable antibacterial activity. This can now also be rationalized on the basis of its large side chains and also the fact that about 60% of its conformers have unrecognizable B2 torsions (data not shown). With regard to the dimensions of the other glutamine analogues, the even larger size of FMDB would be expected to limit severely its transport as part of a peptidomimetic, which would further diminish its poor enzyme inhibition. The small size of FCDP supports the finding that DppA binds FCDP-Ala and FCDP-Met better than FMDP-Ala and FMDP-Met (Table 2), which endorses the suggestion that the poor antibacterial activities of FCDP peptidomimetics are probably a consequence of the intrinsically lower inhibition of the target enzyme by this analogue. Nature endorses this influence of side chain size on bioactivity. Thus, most natural antimicrobial dipeptidomimetics have a protein amino acid as their N-terminus, commonly Ala, and the larger antimicrobial compound forms the C-terminus. This is invariably the case when the inhibitory moiety is large, as in valclavam, polyoxins and nikkomycins.17,22,23,26,27 Interestingly, the natural antibiotic lindenbein, FCDP-Ala, has the warhead at the N-terminus. However, as shown above, being the smallest of these glutamine analogues it can be accommodated in the N-terminal pocket, allowing it to be well bound by DppA (Table 2), although FCDP has poor inhibitory activity against the target enzyme (Table 1). Molecular modelling of Ala-FCDP, an alternative theoretical lindenbein analogue, shows that compared with the natural form it possesses a much lower proportion of conformers that match the MRT for Dpp (results not shown). It appears that nature chose the form best suited to deliver the antimicrobial moiety.
Lys-FMDP provides a further example of the ability of side chains to affect molecular recognition. Its conformers are mostly A4(B9,B12), which are putative ligands specifically for Tpp, but it was completely inactive with this transporter. The explanation for this became clear from inspection of the modelled conformers in which their side chains interact with one another. This may improve stability but would seem to make the fused side chains too large to allow binding by Tpp. With this insight, it would be relatively easy to evaluate conformer databases of related compounds to identify any in which the side chains are comparably close and so may constitute an unrecognizable fused unit.
In conclusion, conformational analysis of these peptidomimetics has provided detailed insights into their structureactivity relationships. This approach would speed up identification of compounds with optimal bioactivities whilst dramatically decreasing the need for extensive chemical syntheses and testing. In addition, molecular modelling could be performed on, for example, FMDP-containing pseudopeptides, having chemical features such as peptide-bond isosteres that could enhance overall pharmacokinetics.34 In a broader context, the study extends the use of the concept of MRTs, and provides important structural information that will aid the design not only of antimicrobial compounds but also of all therapeutic agents targeted for delivery by peptide transporters.
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Acknowledgements |
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Footnotes |
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References |
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