Protonation of phosphoramide mustard and other phosphoramides

Mp, Gamcsik; Sm, Ludeman; Em, Shulman-Roskes; Ij, McLennan; Me, Colvin; Om, Colvin

doi:10.1021/jm00075a019

Cited by 22 publications

(40 citation statements)

References 13 publications

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“…This transition must be the result of the phosphoramide nitrogen deprotonation. Indeed, the magnitude of the induced shift upon titration (13.3 ppm) as well as the direction of it (downfield shift upon ionization) are in accordance with nitrogen deprotonation (33).…”

Section: Roesy) Interactionssupporting

confidence: 54%

Chemical Structure and Translation Inhibition Studies of the Antibiotic Microcin C7

Guijarro

González-Pastor

Baleux

et al. 1995

Journal of Biological Chemistry

129

131

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Microcin C7 (MccC7) 1 is a small peptide antibiotic produced by Escherichia coli during the stationary phase of growth (1). The spectrum of activity of this microcin includes several members of the Enterobacteriaceae family (2). MccC7 exerts its bacteriostatic action by blocking protein synthesis. Like for other microcins, the bacterial strains that produce MccC7 are immune (resistant) to this microcin (3). The heptapeptide of MccC7 is synthesized in ribosomes (4) and undergoes posttranslational modifications to yield the mature molecule.The genetic determinants for MccC7 synthesis, export, and immunity have been cloned from the 43 kilobases E. coli pMccC7 plasmid into multicopy plasmids that overproduce MccC7 and express MccC7 immunity (5). These determinants lie on a 6.2-kilobase region of pMccC7, which has been entirely sequenced.2 Different complementary approaches, such as physical and phenotypical characterization of insertion mutations and complementation studies, have shown that this region contains six genes (mccABCDEF) (5).2 Genes A, B, C, D, and E are involved in the production of mature extracellular microcin. Genes C, E, and F code for self-immunity bestowing products. Genes mccA-D are directly involved in the synthesis and export of MccC7 and constitute an operon transcribed from a promoter (mccp), located upstream of mccA (4, 6). Expression of these genes is regulated by the cAMP-cAMP regulatory protein complex and by the stationary phase factor RpoS (also called AppR) (6, 7). 3 mccA codes for the unmodified peptide of MccC7, MRTGNAN (MccA) (4). The predicted gene polypeptide product of mccB (350 residues) is strikingly homologous to a 81-residue fragment of the ubiquitin-activating enzyme from different eucaryotic species (UBA1) (8), ThiF (9) and ChlN (10) from E. coli which participate, respectively, in the biosynthesis of thiamine pyrophosphate and of molibdopterin, and HesA, an enzyme required by Anabaena for nitrogen fixation (11). The predicted mccC product (404 residues) contains 11 potential transmembrane domains and displays significant similarity with stretches of transport proteins, 2 suggesting that MccC is responsible for MccC7 export and explaining why it also confers resistance to exogenous MccC7. The carboxyl end of the expected MccE 521-residue long polypeptide is highly similar to RimL, an enzyme that acetylates the ribosomal protein L12 from E. coli (12). Principally on the basis of this homology with an acetylating protein, and because target alteration is a common mechanism of antibiotic resistance (13), it has been proposed that MccE might confer MccC7 immunity to producing cells by acetylating the target of microcin C7.2 No similarity was found for the predicted MccD (267 residues) and MccF (334 residues) polypeptides.The knowledge of the chemical structure of MccC7 is neces-* This investigation was supported by funds from the Institut Pasteur, the Centre National de la Recherche Scientifique (URA 1129 and URA 489), and the European Union (Grant CI1*-CT92-0011 to F. M.). The...

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Section: Roesy) Interactionssupporting

confidence: 54%

Chemical Structure and Translation Inhibition Studies of the Antibiotic Microcin C7

Guijarro

González-Pastor

Baleux

et al. 1995

Journal of Biological Chemistry

129

131

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“…No resonances indicative of phosphoramide mustard (4a) were observed during the course of the experiment, as verified by the addition of authentic material to the NMR sample. 29 The chemical shifts and stabilities of the downfield signals (6 19.4 and 19.3) were consistent with those expected for the E and Z isomers of the expected acyclic product, aldophosphamide 0-(2,3,4,5,6-pentafluorobenzyl)oxime (EIZ-5a) Scheme 1).21 Subsequently, the isomers were isolated as a mixture and identified by IH NMR. In the proton spectrum, triplets attributable to the E/Z imino proton (CH=N) were observed at 7.43 and 6.83 ppm in a ratio of 3 2 , respectively.…”

Section: Resultssupporting

confidence: 55%

Oxime Derivatives of the Intermediary Oncostatic Metabolites of Cyclophosphamide and Ifosfamide: Synthesis and Deuterium Labeling for Applications to Metabolite Quantification

Ludeman

Shulman-Roskes

Wong

et al. 1995

Journal of Pharmaceutical Sciences

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“…In serine proteases, the pK a of the catalytic triad histidine side has been determined to be close to 7.0 (21) and consistent with its role in protonating the departing α-NH 2 of the peptide, which would have a pK a of greater than 9.0. However, in the case of a phosphoramide leaving group of McC7, the pK a of the α-NH 2 would be close to 4.5 (22), making protonation by the catalytic histidine side chain (His311) difficult. The McC7 α-NH 2 can only be protonated by either a glutamate or aspartate (pK a lower than 4.5), and no candidate carboxylate is present in the vicinity of the departing amine.…”

Section: Discussionmentioning

confidence: 99%

Structure and function of a serine carboxypeptidase adapted for degradation of the protein synthesis antibiotic microcin C7

Agarwal

Tikhonov

Metlitskaya

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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Several classes of naturally occurring antimicrobials exert their antibiotic activity by specifically targeting aminoacyl-tRNA synthetases, validating these enzymes as drug targets. The aspartyl tRNA synthetase “Trojan horse” inhibitor microcin C7 (McC7) consists of a nonhydrolyzable aspartyl-adenylate conjugated to a hexapeptide carrier that facilitates active import into bacterial cells through an oligopeptide transport system. Subsequent proteolytic processing releases the toxic compound inside the cell. Producing strains of McC7 must protect themselves against autotoxicity that may result from premature processing. The mccF gene confers resistance against endogenous and exogenous McC7 by hydrolyzing the amide bond that connects the peptide and nucleotide moieties of McC7. We present here crystal structures of MccF, in complex with various ligands. The MccF structure is similar to that of dipeptide ld -carboxypeptidase, but with an additional loop proximal to the active site that serves as the primary determinant for recognition of adenylated substrates. Wild-type MccF only hydrolyzes the naturally occurring aspartyl phosphoramidate McC7 and synthetic peptidyl sulfamoyl adenylates that contain anionic side chains. We show that substitutions of two active site MccF residues result in a specificity switch toward aromatic aminoacyl–adenylate substrates. These results suggest how MccF-like enzymes may be used to avert various toxic aminoacyl–adenylates that accumulate during antibiotic biosynthesis or in normal metabolism of the cell.

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Protonation of phosphoramide mustard and other phosphoramides

Cited by 22 publications

References 13 publications

Chemical Structure and Translation Inhibition Studies of the Antibiotic Microcin C7

Chemical Structure and Translation Inhibition Studies of the Antibiotic Microcin C7

Oxime Derivatives of the Intermediary Oncostatic Metabolites of Cyclophosphamide and Ifosfamide: Synthesis and Deuterium Labeling for Applications to Metabolite Quantification

Structure and function of a serine carboxypeptidase adapted for degradation of the protein synthesis antibiotic microcin C7

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