Electrophilic RNA for Peptidyl-RNA Synthesis and Site-Specific Cross-Linking with tRNA-Binding Enzymes.
RNA functionalization is challenging due to the instability of RNA and the limited range of available enzymatic reactions. We developed a strategy based on solid phase synthesis and post-functionalization to introduce an electrophilic site at the 3' end of tRNA analogues. The squarate diester used as an electrophile enabled sequential amidation and provided asymmetric squaramides with high selectivity. The squaramate-RNAs specifically reacted with the lysine of UDP-MurNAc-pentapeptide, a peptidoglycan precursor used by the aminoacyl-transferase FemXWvfor synthesis of the bacterial cell wall. The peptidyl-RNA obtained with squaramate-RNA and unprotected UDP-MurNAc-pentapeptide efficiently inhibited FemXWv. The squaramate unit also promoted specific cross-linking of RNA to the catalytic Lys of FemXWvbut not to related transferases recognizing different aminoacyl-tRNAs. Thus, squaramate-RNAs provide specificity for cross-linking with defined groups in complex biomolecules due to its unique reactivity.
Laboratoire de Recherche Moléculaire sur les Antibiotiques Centre de Recherche des Cordeliers, Equipe 12, UMR S 1138; INSERM, Université Pierre et Marie Curie-Paris 6, Université Paris Descartes, 15 rue de L'Ecole de Médecine, Paris, F-75006, France.
Matthieu
M
Fonvielle
Laboratoire de Chimie et de Biochimie Pharmacologiques et Toxicologiques, Université Paris Descartes, UMR 8601, Paris, F-75006, France.|CNRS UMR 8601, Paris, F-75006, France.
Nicolas
N
Sakkas
Laboratoire de Chimie et de Biochimie Pharmacologiques et Toxicologiques, Université Paris Descartes, UMR 8601, Paris, F-75006, France.|CNRS UMR 8601, Paris, F-75006, France.
Laura
L
Iannazzo
Laboratoire de Recherche Moléculaire sur les Antibiotiques Centre de Recherche des Cordeliers, Equipe 12, UMR S 1138; INSERM, Université Pierre et Marie Curie-Paris 6, Université Paris Descartes, 15 rue de L'Ecole de Médecine, Paris, F-75006, France.
Chloé
C
Le Fournis
Institute for Integrative Biology of the Cell (I2BC), CEA, CNRS, Univ Paris-Sud, Université Paris-Saclay, 91198, Gif-sur-Yvette cedex, France.
Delphine
D
Patin
Institute for Integrative Biology of the Cell (I2BC), CEA, CNRS, Univ Paris-Sud, Université Paris-Saclay, 91198, Gif-sur-Yvette cedex, France.
Dominique
D
Mengin-Lecreulx
Department of Chemistry, University of Oxford, Chemistry Research Laboratory, 12 Mansfield Road, Oxford, OX1 3TA, UK.|Chemistry Branch, Dept. of Science and Mathematics, Faculty of Petroleum and Mining Engineering, Suez Canal University, Suez, 43721, Egypt.
Afaf
A
El-Sagheer
Laboratoire de Chimie et de Biochimie Pharmacologiques et Toxicologiques, Université Paris Descartes, UMR 8601, Paris, F-75006, France.|CNRS UMR 8601, Paris, F-75006, France.
Emmanuelle
E
Braud
Laboratoire de Recherche Moléculaire sur les Antibiotiques Centre de Recherche des Cordeliers, Equipe 12, UMR S 1138; INSERM, Université Pierre et Marie Curie-Paris 6, Université Paris Descartes, 15 rue de L'Ecole de Médecine, Paris, F-75006, France.
Sébastien
S
Cardon
Department of Chemistry, University of Oxford, Chemistry Research Laboratory, 12 Mansfield Road, Oxford, OX1 3TA, UK.
Tom
T
Brown
Laboratoire de Recherche Moléculaire sur les Antibiotiques Centre de Recherche des Cordeliers, Equipe 12, UMR S 1138; INSERM, Université Pierre et Marie Curie-Paris 6, Université Paris Descartes, 15 rue de L'Ecole de Médecine, Paris, F-75006, France. michel.arthur@crc.jussieu.fr.
Michel
M
Arthur
Laboratoire de Chimie et de Biochimie Pharmacologiques et Toxicologiques, Université Paris Descartes, UMR 8601, Paris, F-75006, France. melanie.etheve-quelquejeu@parisdescartes.fr.|CNRS UMR 8601, Paris, F-75006, France. melanie.etheve-quelquejeu@parisdescartes.fr.
Mélanie
0000-0002-4105-3243
M
Etheve-Quelquejeu
D019881:Aminoacyltransferases / Q000737:chemistry / Q000378:metabolism*; D003432:Cross-Linking Reagents / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D008968:Molecular Conformation; D010455:Peptides / Q000737:chemistry / Q000378:metabolism*; D012313:RNA / Q000096:biosynthesis* / Q000737:chemistry; D012343:RNA, Transfer / Q000737:chemistry / Q000378:metabolism*; D014538:Uridine Diphosphate N-Acetylmuramic Acid / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism
D003432:Cross-Linking Reagents; D010455:Peptides; D014538:Uridine Diphosphate N-Acetylmuramic Acid; C002225:UDP-N-acetylmuramic acid pentapeptide; D012313:RNA; D012343:RNA, Transfer; D019881:Aminoacyltransferases
Fem transferases; RNA modifications; crosslinking; post-functionalization; squarates