Negative mode MS/MS to read digital information encoded in sequence-defined oligo(urethane)s: A mechanistic study

International audienceMS/MS sequencing is an unrivaled technique to decipher binary information chemically encoded in the backbone of sequence-controlled synthetic polymers constructed with two co-monomers of different mass, arbitrarily designated as the 0- and 1-bit of the ASCII alphabet. Efficiency of this "reading" step relies however on the simplicity of MS/MS patterns, which depends on both polymer chemistry and chain length. In this context, polyurethanes (PUs) were very promising candidates as dissociation of small deprotonated oligomers (n \\textless 8) yielded a single fragment series. The carbamate bond cleavage reaction was hence studied in details to tentatively anticipate the CID behavior of longer chains prior to optimizing their synthesis. In spite of the simplicity of MS/MS spectra, three different mechanisms were evidenced; however, they were not expected to induce MS/MS complexity when activating longer chains, as verified for sequence-controlled PUs containing up to two bytes of information (i.e., 16 co-monomers). In contrast, the ionization step appeared to be an issue: deprotonation yield of the end-group in negative ion mode electrospray was observed to strongly decrease as PU chain length increases. This sensitivity issue was addressed by introducing a second acidic end-group to allow doubly deprotonated oligomers with no impact on their CID behavior. (C) 2017 Elsevier B.V. All rights reserved

Méthodologies de séquençage MS/MS de polymères encodés

L’objectif de ces travaux de thèse était de développer des stratégies de séquençage MS/MS pour lire des messages binaires écrits dans des polymères synthétiques grâce à une séquence définie de deux co-monomères. Lire de tels messages requiert une couverture de séquence totale, obtenue lorsque les ruptures du squelette polymérique génèrent un nombre limité de séries de fragments. Parce que le comportement dissociatif d’un polymère est dicté par la nature de ses liaisons chimiques, la stratégie mise en œuvre a consisté à utiliser les données MS/MS des chaînes courtes pour optimiser la structure de longs polymères, comme appliqué au cas des poly(phosphodiester)s difficiles à séquencer au delà de DP 50 du fait de la dissociation complexe des groupements phosphates. Leur structure a été modifiée en plaçant des liaisons alcoxyamines entre chaque unité (ou bit) pour rendre toute autre liaison muette en MS/MS. La résolution instrumentale a limité le séquençage à des chaînes de DP 40, ce qui correspond à 80 bits d’information lorsque l’alcoxyamine est utilisée comme un second segment codant pour augmenter la densité de stockage. Un second design moléculaire encore plus efficient a conduit aux poly(alcoxyamine phosphodiester)s à blocs, dans lesquels une liaison alcoxyamine est placée entre chaque octet d’information. La lecture de longs messages est réalisée par MS3, où les fragments générés pendant la première étape d’activation contiennent un seul octet et sont séquencés individuellement au cours de la seconde étape. L’utilisation d’un système de marqueurs pour déterminer la localisation initiale de chaque octet permet de restituer la séquence complèteThis thesis work aimed at developing MS/MS sequencing strategies to read binary messages written in synthetic polymers based on a defined sequence of two co-monomers. Reading such messages requires full sequence coverage, which can be achieved when dissociation reactions occur via efficient backbone bond cleavages but yield a limited number of fragment series. Because dissociation behavior of polymers is dictated by the backbone chemistry, the strategy developed here was to use MS/MS data to re-engineer the structure of long polymers, as applied here to poly(phosphodiester)s, hardly sequenced above DP 50 due to complex dissociation of phosphate groups. Their structure was modified by placing low dissociation energy alkoxyamine linkages between each unit (or bit), which made any other chemical bonds MS/MS-silent. Due to instrumental resolution capabilities, the upper sequencing limit was DP 40, corresponding to 80 bits of information when using the alkoxyamine moiety as a second coding segment to increase storage density. This first design paved the way to the development of byte-truncated poly(alkoxyamine phosphodiester)s, a more MS/MS-efficient structure in which alkoxyamine bonds were placed between each byte of information. Long messages could be read in MS3 experiments, where single-byte fragments released during the first activation stage were further dissociated for sequencing. The whole sequence was re-constructed thanks to a mass tagging system used to determine the original location of each byte in the polymer chai

MS/MS Digital Readout: Analysis of Binary Information Encoded in the Monomer Sequences of Poly(triazole amide)s

International audienceTandem mass spectrometry was evaluated as a reliable sequencing methodology to read codes encrypted in monodisperse sequence-coded oligo(triazole amide)s. The studied oligomers were composed of monomers containing a triazole ring, a short ethylene oxide segment, and an amide group as well as a short alkyl chain (propyl or isobutyl) which defined the 0/1 molecular binary code. Using electrospray ionization, oligo(triazole amide)s were best ionized as protonated molecules and were observed to adopt a single charge state, suggesting that adducted protons were located on every other monomer unit. Upon collisional activation, cleavages of the amide bond and of one ether bond were observed to proceed in each monomer, yielding two sets of complementary product ions. Distribution of protons over the precursor structure was found to remain unchanged upon activation, allowing charge state to be anticipated for product ions in the four series and hence facilitating their assignment for a straightforward characterization of any encoded oligo(triazole amide)s

Mass spectrometry sequencing of long digital polymers facilitated by programmed inter-byte fragmentation

Digital information can be stored in monomer sequences of non-natural macromolecules, but only short chains can be read. Here the authors show long multi-byte digital polymers sequenced in a moderate resolution mass spectrometer. Full sequence coverage can be attained without pre-analysis digestion or the help from sequence databases

Revealing data encrypted in sequence-controlled poly(alkoxyamine phosphodiester)s by combining ion mobility with tandem mass spectrometry

International audienc

Universal Soluble Polymer Supports with Precisely Controlled Loading Capacity for Sequence-defined Oligomer Synthesis

International audienceSoluble supports, with optimal molecular structures for iterative oligomer synthesis, were prepared by atom transfer radical copolymerization of styrene with Fmoc-amino ethyl methacrylate. Size exclusion chromatography and nuclear magnetic resonance indicated formation of copolymers with controlled chain lengths, chain ends, dispersity, and comonomer compositions. These polymers were afterward subjected to two subsequent modifications steps: (a) the debromination of their ω-chain ends via a tributyltin hydride treatment and (b) theremoval of the Fmoc-protecting groups in acidic conditions, thus leading to bromine-free copolymers with a precise primaryamine loading capacity. These universal amine-containing precursors were then derived into a variety of functional supports. A glycine-loaded Wang-type soluble polystyrene support was prepared in two steps and tested for peptide synthesis as well as for the preparation of digital oligo(alkoxyamine amide)s. In both cases, it was possible to obtain uniform sequence-defined oligomers as evidenced by electrospray ionization tandem mass spectrometry. However, each type of oligomer required specific experimental conditions and therefore soluble supportswith an adapted loading capacity for optimal synthesis. Furthermore, the amine-containing universal precursors were also transformed into soluble supports for iterative phosphoramidite chemistry. A support was specifically conceived for the iterative synthesis of abiotic digital oligo(phosphodiester)s and another one was designed for oligonucleotide synthesis

Desorption Electrospray Ionization (DESI) of Digital Polymers: Direct Tandem Mass Spectrometry Decoding and Imaging from Materials Surfaces

International audienceSequence-defined synthetic polymers have recently emerged as an attractive medium to store information at the molecular level, where comonomers of the chains are defined as letters of an alphabet. The main read-out methodology employed to retrieve such molecularly encoded information is tandem mass spectrometry (MS/MS), but a major current limitation remains the low storage capacity of readable chains. Ordering short oligomers at discrete locations onto surfaces to compose long messages is an attractive alternative to the difficult synthesis of long coded polymers. Yet, such surface storage requires a reliable sampling technique to be coupled on-line with MS/MS. Because it combines fast surface extraction with efficient analyte ionization in ambient conditions, desorption electrospray ionization (DESI) is shown here to be perfectly suited to envisage bidimensional data storage. The present study demonstrates performances of DESI-MS/MS at mapping oligomers used to write letters of a word, extracting digital labels from materials tagged for anticounterfeiting purposes, and imaging text written with coded polymeric inks

Cleavable Binary Dyads: Simplifying Data Extraction and Increasing Storage Density in Digital Polymers

WOS:000432710100051Digital polymers are uniform macromolecules that store monomer-based binary sequences. Molecularly stored information is usually extracted from the polymer by a tandem mass spectrometry (MS/MS) measurement, in which the coded chains are fragmented to reveal each bit (i.e. basic coded monomer unit) of the sequence. Here, we show that data-extraction can be greatly simplified by favoring the formation of MS/MS fragments containing two bits instead of one. In order to do so, digital poly(alkoxyamine phosphodiester)s, containing binary dyads in each repeat unit, were prepared by an orthogonal solid-phase approach involving successive phosphoramidite and radical-radical coupling steps. Three different sets of monomers were considered to build these polymers. In all cases, four coded building blockstwo hydroxy-nitroxides and two phosphoramidite monomerswere required to build the dyads. Among the three studied monomer sets, one combination allowed synthesis of unifor m sequence-coded polymers. The resulting polymers led to clear dyad-containing fragments in MS/MS and could therefore be efficiently decoded. Additionally, an algorithm was created to detect specific dyad fragments, thus enabling automated sequencing

Indirect Tertiary Alcohol Enantiocontrol by Acylative Organocatalytic Kinetic Resolution

The stereocontrol of tertiary alcohols represents a recurrent challenge in organic synthesis. In the present paper, we describe a simple, efficient, and indirect method to enantioselectively prepare tertiary alcohols through a chiral isothiourea catalyzed selective acylation of adjacent secondary alcohols. This transformation enables the kinetic resolution (KR) of easily prepared racemic diastereoenriched secondary/tertiary diols providing both monoesters and starting diols in highly enantioenriched forms (s-value >200)